bluuebunny/crossref_metadata_2025 huggingface dataset:
Improved Motor Function of Denervated Rat Hindlimb Muscles Induced by Embryonic Spinal Cord Grafts
Authors: Nógrádi Antal , Vrbová Gerta
Published: April 2006

Loss of motoneurons results in a decrease in force production by skeletal muscles and paralysis. Although it has been shown that missing motoneurons of rats can be replaced by embryonic homotopic neurons, attempts to guide their axons to their target muscles that have lost their innervation have been unsuccessful. In this study attempts were made to guide axons from grafted embryonic motoneurons to their target via a reimplanted ventral root. Adult hosts that received an embryonic graft prelabelled with 5‐bromo‐2′‐deoxyuridine had their L4 ventral root avulsed and reimplanted into the spinal cord. Three to six months later, neurons that had their axons in the L4 ventral ramus were retrogradely labelled with fast blue and diamidino yellow. In five animals that had received an embryonic graft 116 ± 16 cells were retrogradely labelled, and of these at least 15% were of graft origin, since they were positive for 5‐bromo‐2′‐deoxyuridine. In five animals that had their L4 ventral root reimplanted but did not receive a graft, only 12 ± 1.3 cells were retrogradely labelled. However, meaningful functional recovery could be achieved only if the regenerating axons of embryonic motoneurons found in the L4 ventral ramus were able to reverse the loss of force of muscles that had lost their innervation. This study shows that axons of embryonic motoneurons grafted into an adult rat spinal cord, as well as some axons of host origin, can be guided to denervated hindlimb muscles via reimplanted lumbar ventral roots. In normal rats ∼30 motor axons innervated the extensor digitorurn longus and 60 innervated the tibialis anterior via the L4 ventral root. In rats that did not receive a graft only 3.7 ± 1.2 axons reached the extensor digitorum longus and 3.5 ± 0.4 reached the tibialis anterior muscle via the implanted L4 ventral root. In animals that had an embryonic graft, 7.6 ± 0.5 axons innervated the extensor digitorum longus and 8.5 ± 0.5 reached the tibialis anterior muscle via the implanted root. In rats without a transplant the maximum tetanic tension elicited by stimulating the implanted L4 root was 16 ± 7 g for the extensor digitorum longus and 53 ± 36 g for the tibialis anterior muscle, whereas the corresponding muscles in animals that had an embryonic graft developed 82 ± 16 and 281 ± 95 g respectively. Thus it appears that the grafted motoneurons contributed to the innervation and functional recovery of the denervated muscles.
Postnatal Changes in the Expression of the trkA High‐affinity NGF Receptor in Primary Sensory Neurons
Authors: Ennett David L. H. , Averill Sharon , Clary Douglas O. , Priestley John V. , McMahon Stephen B.
Published: April 2006

In development ∼70–80% of dorsal root ganglion (DRG) cells are dependent on nerve growth factor (NGF) for their survival, while in the adult only some 40% of DRG cells express the high‐affinity NGF receptor, trkA. This discrepancy suggests that trkA expression, and therefore neurotrophin sensitivity, may alter as the animal matures. We have tested this possibility by counting the number of L4/5 DRG neurons showing immunoreactivity for trkA in rats from the day of birth to postnatal day 14. We also examined changes in p75 and IB4 labelling. On the day of birth, 71% of DRG cells were found to express trkA. However, this percentage gradually fell with age and reached adult levels at postnatal day 14. The expression of p75 did not parallel that of trkA, remaining relatively constant at between 45 and 50% of cells from birth to postnatal day 14. Over the same period there was a marked increase in the proportion of cells which bind the lectin IB4 from 9 (day of birth) to 40% (day 14). Since in the adult the 1B4 population consists of small cells which mostly do not express trkA, this finding suggests that the postnatal down‐regulation of trkA occurs in this population. Consistent with this suggestion are the results of double labelling for trkA and IB4, which confirmed that at times intermediate between birth and postnatal day 14 there was a high degree of coexpression between these markers (which is absent in the adult). This result also suggests that the down‐regulation of trkA is unlikely to be directly responsible for the emerging IB4 binding.
C‐fos Expression During Vocal Mobbing in the New World Monkey Saguinus fuscicollis
Authors: Jürgens U. , Lu C.‐L. , Quondamatteo F.
Published: April 2006

In order to find brain areas involved in the vocal expression of emotion, we compared c‐fos expression in three groups of saddle‐back tamarins (Saguinus fuscicollis). One group, consisting of three animals, was made to utter more than 800 mobbing calls by electrical stimulation of the periaqueductal grey of the midbrain (PAG). A second group, consisting of two animals, was stimulated in the PAG with the same intensity and for the same duration as the first group but at sites that did not produce vocalization. These sites lay somewhat medial to the vocalization‐eliciting sites. A third group, consisting of two animals, was stimulated at vocalization‐eliciting sites in the PAG but with an intensity below vocalization threshold. Fos‐like immunoreactivity that was found in the vocalizing but not in the non‐vocalizing animals was located in the dorsomedial and ventrolateral prefrontal cortex, anterior cingulate cortex, ventrolateral premotor cortex, sensorimotor face cortex, insula, inferior parietal cortex, superior temporal cortex, claustrum, entorhinal and parahippocampal cortex, basal amygdaloid nucleus, anterior and dorsomedial hypothalamus, nucleus reuniens, lateral habenula, Edinger‐Westphal nucleus, ventral and dorsolateral midbrain tegmentum, nucleus cuneiformis, sagulum, pedunculopontine and laterodorsal tegmental nuclei, ventral raphe, periambigual reticular formation and solitary tract nucleus. For some of these structures (e.g. anterior cingulate cortex and periambigual reticular formation), there is evidence also from electrical stimulation, lesioning and single‐unit recording studies that they are involved in vocal control. For other structures (e.g. lateral habenula, Edinger‐Westphal nucleus), the available evidence speaks against such a role. Fos activation in these cases is probably related to non‐vocal reactions accompanying the electrically elicited vocalizations. A third group of structures consists of areas for which a role in vocal control cannot be excluded but for which the present study presents the first evidence for such a role (e.g. claustrum and sagulum). These structures deserve further studies using more specific methods.
Accurate Synapse Regeneration Despite Ablation of the Distal Axon Segment
Authors: Mason Adrian , Muller Kenneth J.
Published: April 2006

In each body ganglion of the leech Hirudo medicinalis there is a single S‐cell. After an S‐cell axon is severed, it regenerates along its surviving distal segment and reconnects with its synaptic target, the axon of the neighbouring S‐cell. In approximately half the cases the regenerating axon forms a temporary electrical synapse specifically with the distal segment, which remains active and connected to the target, thereby functioning as a splice until regeneration is complete. To determine whether the distal axon segment is required for successful regeneration, distal segments of severed S‐cell axons were ablated by intracellular injection of bacterial protease. Fifty‐seven preparations were examined from 2 to 212 days after injection of the axon segment. The extent of S‐cell axon regeneration was assessed electrophysiologically by intracellular and extracellular recording, and anatomically by intracellular injection of markers followed by light microscopy and electron microscopy. The S‐cell axons regenerated successfully in almost 90% of animals examined after 2 weeks or more. In a further four animals the target S‐cell was ablated in addition to the distal axon segment, permanently disrupting conduction along the S‐cell pathway. Nevertheless, the regenerating axon grew along its usual pathway and there was no evidence that alternative connections were formed. It is concluded that, although the distal axon segment can provide a means for rapid functional repair, the segment is not required for reliable regeneration of the axon along its usual pathway and accurate formation of an electrical synapse.
Ontogenetic Development of 5‐HT1D Receptors in Human Brain: An Autoradiographic Study
Authors: del Olmo Elena , del Arco Carmen , Díaz Alvaro , Pascual Julio , Mengod Guadalupe , Palacios José M. , Pazos Angel
Published: April 2006

The pattern of pre‐ and postnatal appearance of 5‐HT1D receptors throughout the different areas of the human brain was studied by quantitative in vitro autoradiography, using [125I]GTI (serotonin O‐carboxymethyl‐glycyl‐[125I]tyrosinamide) as a ligand. The anatomical distribution of 5‐HT1D receptors in neonatal, infant and children's brain was in good agreement with that observed in the adult, the basal ganglia and substantia nigra being the most intensely labelled areas. The development of these receptors throughout the human brain was mainly postnatal: low densities of [125I]GTI binding sites were observed at the fetal/neonatal stage in most regions analyzed, in contrast with the high levels of labelling found in infant and children's brains. Indeed, in a number of regions, including the globus pallidus, substantia nigra and visual cortex, a peak of overexpression of 5‐HT1D receptors was observed in the first decade of life. Such overexpression could support a regulatory role for 5‐HT1D receptors in advanced periods of the CNS developmental process. Our results also indicate that the administration of drugs acting on 5‐HT1D receptors during the early postnatal period of life could result in modifications of their properties, as these receptors are already functional in this period.
Further Observations on the Role of Nitric Oxide in the Feline Lateral Geniculate Nucleus
Authors: Cudeiro J. , Rivadulla C. , Rodriguez R. , Martinez‐Conde S. , Martinez L. , Grieve K. L. , Acuña C.
Published: April 2006

We have examined the responses of a population of 77 cells in the dorsal lateral geniculate nucleus (dLGN) of the anaesthetized, paralysed cat. Here the synthetic enzyme for the production of nitric oxide, nitric oxide synthase, is found only in the presynaptic terminals of the cholinergic input from the brainstem. In our hands, iontophoretic application of inhibitors of this enzyme resulted both in significant decreases in visual responses and decreased responses to exogenous application of NMDA, effects which were reversed by coapplication of the natural substrate for nitric oxide synthase, L‐arginine, but not the biologically inactive isomer, D‐arginine. Nitroprusside and S‐nitroso‐N‐acetylpenicillamine (SNAP), nitric oxide donors, but not L‐arginine, were able to increase markedly both spontaneous activity and the responsiveness to NMDA application. Furthermore, SNAP application facilitated visual responses. Responses of cells in animals without retinal, cortical and parabrachial input to the LGN suggest a postsynaptic site of action of nitric oxide. This modulation of the gain of visual signals transmitted to the cortex suggests a completely novel pathway for nitric oxide regulation of function, as yet described only in primary sensory thalamus of the mammalian central nervous system.
Neurotensin Excitation of Serotonergic Neurons in the Dorsal Raphe Nucleus of the Rat In Vitro
Authors: Jolas Thierry , Aghajanian George K.
Published: April 2006

Neurotensin‐containing terminals and radioligand binding sites are present in the dorsal raphe nucleus. The purpose of this study was to test, in brain slices containing this nucleus, the effect of neurotensin on the electrical activity of serotonergic neurons. In extracellular recordings, the cells were identified by the ability of the α1‐adrenoceptor agonist phenylephrine to induce firing, and serotonin to reduce this effect. After washout of phenylephrine, neurotensin (10 nM to 10 μM) induced a concentration‐dependent increase in the firing rate of serotonergic neurons (EC50= 142 nM; maximum effect ˜1 μM). The neurotensin excitation, which was mimicked by neurotensin fragments 8–13 but not neurotensin peptide fragment 1–8 and selectively blocked by SR 48692 (100 nM), was observed mainly in the ventral part of the nucleus. Most serotonergic neurons showed marked desensitization to neurotensin, even at low concentrations. The neurotensin response was occluded by supramaximal concentrations of phenylephrine. In intracellular recordings using KCl‐containing electrodes, neurotensin induced an inward current associated in some cases with a decrease in apparent input conductance. In conclusion, neurotensin was found to have an excitatory action on serotonergic neurons in the ventral part of the dorsal raphe nucleus, an effect which could be subject to desensitization and was occluded by phenylephrine. This occlusion phenomenon may be important for the physiological role of neurotensin in the dorsal raphe nucleus.
Differential Oscillatory Properties of Cholinergic and Non‐cholinergic Nucleus Basalis Neurons in Guinea Pig Brain Slice
Authors: Alonso Angel , Khateb Asaid , Fort Patrice , Jones Barbara E. , Mühlethaler Michel
Published: April 2006

Evidence has suggested that the nucleus basalis magnocellularis has the potential to influence the functional state of the cerebral cortex through topographically organized, widespread projections of the cholinergic cells in that nucleus. It has also been shown that, in addition to the cholinergic neurons, other non‐cholinergic magnocellular basal forebrain neurons, some of which have been identified as gamma‐aminobutyric acid‐ergic, project into the cerebral cortex and thus may also participate in the modulation of its activity. We have performed a comparative study of the intrinsic rhythmic properties of immunohistochemically and morphologically characterized choline acetyltransferase (ChAT)‐positive and ChAT‐negative cells of the nucleus basalis by means of intracellular recordings in guinea pig brain slices. Our results demonstrate that relatively large, multipolar cholinergic and non‐cholinergic neurons each display differential voltage‐dependent properties that allow them to discharge rhythmically in spike bursts and spike clusters, respectively, at low frequencies (<10 Hz). Cholinergic cells display bursts of 2–4 action potentials (at ‐200 Hz) riding on low‐threshold spikes recurring at a low frequency (<5 Hz) when depolarized from a membrane potential more negative than ‐55 mV and display low‐frequency (<10–15 Hz) tonic firing when depolarized from a more positive level. In contrast, non‐cholinergic cells fire in a unique mode, displaying non‐adapting clusters of spikes interspersed with rhythmic subthreshold membrane‐potential oscillations when depolarized from levels less negative than ‐55 mV. The spike clusters repeat rhythmically at relatively low frequencies (2–10 Hz). The intracluster spiking frequency is relatively high and coincides approximately with that of the intervening membrane‐potential oscillations (‐20–70 Hz). The cluster frequency of the non‐cholinergic cells corresponds, in the same manner as the burst frequency of the cholinergic cells, to a delta (1–4 Hz) or theta (4–10 Hz) range of activity, whereas the intra‐cluster and tonic spike frequencies of the non‐cholinergic cells correspond to high beta to gamma ranges of electroencephalographic activity (19–30 Hz and 30–60 Hz, respectively). We propose that the different modes of oscillatory firing by the cholinergic and non‐cholinergic basal forebrain cell populations could collectively contribute to the rhythmic modulation of slow and fast rhythms within the cerebral cortex.
Unique Distribution of Somatostatin‐immunoreactive Cells in the Retina of the Tree Shrew (Tupaia belangeri)
Authors: Engelmann Ralf , Peichl Leo
Published: April 2006

Somatostatin‐like immunoreactive cells in the tree shrew retina were studied with the monoclonal antibody S8 against the neuropeptide somatostatin 14. As in some other mammals, immunoreactive somata are exclusively found in the ganglion cell layer. Immunoreactive processes form a sparse main plexus in the inner plexiform layer near the border of the inner nuclear layer; fewer additional processes are found closer to the ganglion cell layer. With retrograde labelling of retinal ganglion cells by injections of the tracer Fast Blue into the superior colliculus and lateral geniculate body and counterstaining of the retinae with S8, ˜5% of the immunoreactive somata were double‐labelled at any retinal location. The vast majority of somatostatin‐like immunoreactive cells are thus displaced amacrine cells. Their somata are distributed over the entire retina. Their population density is highest in the temporal retina, with peak densities of ˜5000 cells/mm2 near the central area and a steep density gradient. In the remaining retina densities are 200–400 cells/mm2, falling to ˜100 cells/mm2 at the retinal margins. This is in stark contrast to the somatostatin‐like immunoreactive cells in other mammalian retinae which have densities of 10–40 cells/mm2 and are confined to restricted retinal regions (inferior retina and/or retinal margin).
Copper Modulation of NMDA Responses in Mouse and Rat Cultured Hippocampal Neurons
Authors: Vlachová Viktorie , Zemková Hana , Vyklický Ladislav
Published: April 2006

The effect of Cu2+ on NMDA receptors was studied in cultured mouse and rat hippocampal neurons using whole‐cell patch‐clamp and a fast perfusion system. Analysis of the Cu2+ concentration‐response curve for inhibition of NMDA‐induced currents suggests that free Cu2+ directly inhibits NMDA receptors with an IC50 of 0.27 μM. Cu2+ was ineffective in blocking NMDA receptor activity when complexed with NMDA or glycine; NMDA‐Cu2+ and glycine‐Cu2+ complexes acted as agonists of similar potency to the free amino acids. The inhibition by Cu2+ (10–100 μM) of responses to 10 μM NMDA was essentially voltage‐independent. The onset of inhibition by 100 μM Cu2+ of responses to 2 FM glutamate acting at NMDA receptors was significantly faster than NMDA receptor deactivation evoked by a sudden decrease in the concentration of glycine or glutamate, or of both agonists. This suggests that CU2+ acts as a non‐competitive antagonist, and does not directly interfere with the binding of glutamate or glycine to their recognition sites on the NMDA receptor complex. In the absence of NMDA the apparent association rate constant for binding of Cu2+ to NMDA receptors, calculated from the rate of onset of block by Cu2+ of test responses to NMDA, was 19 times slower than in the presence of 30 μM NMDA, suggesting that Cuz+ interacts preferentially with agonist‐bound receptors. Our results show that Cu2+ is a potent inhibitor of NMDA receptor‐mediated responses.
Immunohistochemical Localization of Dopamine D Receptors in Rat Retina
Authors: Veruki Margaret L. , Wässle Heinz
Published: April 2006

We have localized the dopamine D1 receptor in rat retina using a subtype‐specific monoclonal antibody. Immunolabelling can be detected in the inner and outer plexiform layers and in a number of cells in the inner nuclear layer. In the inner plexiform layer, labelled processes form four distinct horizontal bands and a series of patches. In order further to characterize the labelling pattern of the D1 receptor antibody, double‐labelling experiments were performed with antibodies against population‐specific neuronal markers in the retina. Antibodies against tyrosine hydroxylase. choline acetyltransferase, calretinin, calbindin, the glutamate transporter GLT‐I, protein kinase C, recoverin and parvalbumin were co‐applied with the D, receptor antibody. With these cell markers we demonstrate that horizontal cells, at least three types of cone bipolar cells and a small number of amacrine cells are immunolabelled for the D1 receptor. In the inner plexiform layer, processes labelled by the D1 receptor antibody are co‐stratified with processes labelled by the GLT‐1 antibody. D1 receptor‐labelled processes are not co‐localized with the processes of amacrine cells and ganglion cells labelled by antibodies against tyrosine hydroxylase, choline acetyltransferase or calretinin. Our results indicate that dopamine Dl receptors are localized predominantly to horizontal cells and cone bipolar cells. Furthermore, the spatial disparity between dopaminergic processes and the site of the majority of D1 receptors supports the idea that in the retina dopamine acts as a neuromodulator that diffuses through extracellular space. The localization of D1 receptors to a number of identified cell types enables future physiological work to be directed towards specific synaptic circuits within the retina.
Cholera and Pertussis Toxins Reveal Multiple Regulation of cAMP Levels in the Rabbit Carotid Body
Authors: Cachero T. G. , Rigual R. , Rocher A. , Gonzalez C.
Published: April 2006

It is known that hypoxia (PO2∼66–18 mm Hg), acting via unknown receptors, increases carotid body cAMP levels in Ca2+‐free solutions, indicating that low PO2 activates adenylate cyclases independently of the action of the released neurotransmitters. The aim of the present work was to investigate the involvement of G proteins in the genesis of the basal level of cAMP and on the increase in cAMP induced by low P02. In carotid body homogenates, cholera toxin‐ and pertussis toxin‐induced [32P] ADP‐ribosylation of two protein bands of =42 and 45 kDa, and =39 and 40 kDa respectively; in both cases, prior incubation of the carotid bodies with the toxins reduced [32P] ADP‐ribosylation by >90%. In intact carotid bodies, cholera toxin treatment increased cAMP levels more in normoxic than in hypoxic organs, indicating that hypoxia releases neurotransmitters acting on receptors negatively coupled to adenylate cyclases. Cholera toxin‐treated carotid bodies incubated in Ca2+‐free solution had identical cAMP levels in normoxia and in hypoxia. In pertussis toxin‐treated normoxic carotid bodies the cAMP level was close to control, but in pertussis toxin‐treated hypoxic carotid bodies cAMP rose to a level similar to those seen in normoxic cholera toxin‐treated organs, indicating that low PO2 releases neurotransmitters acting on receptors positively coupled to adenylate cyclases. Pertussis toxin‐treated carotid bodies incubated in Ca2+‐free solution lost their capacity to increase cAMP in response to hypoxia, indicating that a G protein sensitive to pertussis toxin is needed for this response. This implies that the carotid bodies express a pertussis toxin‐sensitive G protein positively coupled to adenylate cyclases, or that a Gs protein requiring the cooperative action of Go/Gi donated by subunits mediates the increase in cAMP level produced by hypoxia.
Colocalization of Oestrogen Receptor lmmunoreactivity and Preproenkephalin mRNA Expression to Neurons in the Superficial Laminae of the Spinal and Medullary Dorsal Horn of Rats
Authors: Amandusson Åsa , Hermanson Ola , Blomqvist Anders
Published: April 2006

A double‐labelling procedure combining immunohistochemical staining with in situ hybridization using a radiolabelled cRNA probe was employed to demonstrate oestrogen receptor‐like immunoreactivity and preproenkephalin‐A mRNA in the medullary and spinal dorsal horn of female rats. Both markers labelled large numbers of neurons in the substantia gelatinosa and its trigeminal homologue. Many of these neurons were double‐labelled, displaying both oestrogen receptor‐like‐ immunoreactivity and preproenkephalin‐A mRNA; cell counts showed that 40–60% of the of the oestrogen receptor‐like‐immunoreactive cells in the superficial laminae also were labelled for preproenkephalin‐A mRNA, and that 60–70% of the preproenkephalin‐A mRNA‐labelled neurons in the same laminae displayed oestrogen receptor‐like immunoreactivity. Previous studies have shown that oestrogen receptors can bind to the promoter region of the preproenkephalin‐A gene, and studies on the hypothalamus have demonstrated that oestrogen regulates enkephalin expression in select neuronal populations. The present results demonstrate that enkephalinergic neurons in the superficial dorsal horn contain oestrogen receptors and suggest that oestrogen may play an important role in the modulation of sensory and nociceptive processing in the lower medulla and spinal cord.
Macrophage Inflammatory Protein‐1β (MIP‐1β) Produced Endogenously in Brain During E. coli Fever in Rats
Authors: Miñano Francisco J. , Fernández‐Alonso Antonio , Benamar Khalid , Myers R. D. , Sancibrián Manuel , Ruiz Rosa M. , Armengol José A.
Published: April 2006

Macrophage inflammatory protein‐1 (MIP‐1) evokes an intense fever, independent of a prostaglandin mechanism, and is now thought to play an important role in the defence response to bacterial pyrogens. The purpose of this study was 2‐fold: (i) to determine whether the potent doublet of this cytokine, MIP‐1β, is actually produced in the brain in response to a pyrogenic dose of a lipopolysaccharide of Escherichia coli and (ii) to determine the anatomical site of synthesis of this cytokine in the brain. Following the intense fever produced by intraperitoneal administration of lipopolysaccharide in the unrestrained rat, MIP‐1β immunoreactivity was identified post mortem in two regions of the brain implicated in fever: the organum vasculosum laminae terminalis (OVLT) and the anterior hypothalamic, preoptic area (AH/POA). Microinjection of goat anti‐mouse MIP‐1β antibody (anti‐MIP‐1β) directly into the AH/POA markedly suppressed fever in rats in response to lipopolysaccharide. Further, anti‐MIP‐1β administered 180 min after the injection of lipopolysaccharide acted as an antipyretic and reversed the fever induced by the endotoxin. Anti‐MIP‐1β or control immunoglobulin G antibody microinjected into the hypothalamus immediately before the intraperitoneal injection of the control saline did not alter the temperature of the rats. Taken together, the present results demonstrate that MIP‐1β is produced in the brain in response to a bacterial endotoxin. These observations, in the light of earlier data on fever induced by MIP‐1β, further support the hypothesis that endogenously synthesized MIP‐1β acts as an intermediary factor in the evocation of fever by acting on the thermosensitive cells of the brain.
Differential Effects of an L‐type Ca2+ Channel Antagonist on Activity‐ and Phosphorylation‐enhanced Release of Acetylcholine at the Neuromuscular Junction of the Frog In Vitro
Authors: Arenson M. S. , Gill D. S.
Published: April 2006

The effects of the selective L‐type Ca2+‐channel antagonist nimodipine on changes in the electrophysiological correlates of acetylcholine release induced by activity in the motor nerve or by inhibition of protein dephosphorylation were studied in the isolated sartorius muscle of the frog. Nimodipine (1 μM) had no effect on basal miniature endplate potential (mEPP) frequency or on the quantal content of the endplate potential (EPP) evoked at 0.33 Hz. Stimulation of the motor nerve at frequencies of 20, 40 and 50 Hz progressively increased the quantal content and at 50 Hz caused an increase in mEPP frequency measured at the end of the train. Nimodipine (1 μM) had no effect on the change in either parameter caused by activity in the motor nerve. The L‐channel agonist BAYK 8644 had inconsistent effects, causing at 0.33 Hz an increase in quantal content at 55% of the neuromuscular junctions examined. Application of the phosphatase inhibitor okadaic acid (1 μM) caused an increase in miniature endplate potential and/or current [mEPP(C)] frequency but not in quantal content. The increase in spontaneous frequency was reduced by the Ca2+‐channel blockers nimodipine (1 μM) and cadmium (75 μM). The amplitude of mEPP(C)s was increased by okadaic acid (1 μM), but neither the decay time constant of miniature endplate currents (mEPC) nor the amplitude of endplate currents evoked by iontophoretic application of carbachol was so altered. The activity of electric eel acetylcholinesterase was unchanged by okadaic acid. The present data do not support the concept that the recruitment of normally silent L‐type Ca2+‐channels contributes to activity‐dependent increases in acetylcholine release. The results obtained with okadaic acid suggest that protein phosphorylation and dephosphorylation may regulate the activity of L‐type channels and the packaging of acetylcholine.
Correlations between Water Maze Performance and Cortical Somatostatin mRNA and High‐affinity Binding Sites During Ageing in Rats
Authors: Dournaud Pascal , Jazat‐Poindessous Frédérique , Slama Annie , Lamour Yvon , Epelbaum Jacques
Published: April 2006

Somatostatin levels and high‐affinity (somatostatin‐1) binding sites are decreased in post‐mortem cortical samples of Alzheimer's disease patients but the relationships between such modifications and the cognitive deficits remain to be established. We investigated these relationships in the ageing rat. Three age groups (3–4, 14–15 and 26–27 months) were tested in a modified version of the Morris water maze. Somatostatin mRNA levels were quantified by in situ hybridization and somatostatin binding sites by radioautography using the selective agonist octreotide (SMS 201995) as a competing drug to evaluate high‐affinity (somatostatin‐1) and low‐affinity (somatostatin‐2) binding sites. The number of somatostatin mRNA‐containing cells was not modified with age or memory performance in cortical, hippocampal and hypothalamic regions, but somatostatin mRNA densities were significantly decreased with age and with memory performance in the frontal and parietal cortex. In the frontal cortex somatostatin mRNA densities were already decreased in 14‐ to 15‐month‐old rats, whereas the decrease was observed only in 26‐ to 27‐month‐old rats in the parietal cortex. A decrease in somatostatin‐1 binding was observed with memory performance, independently of age, in the basolateral amygdala only, while somatostatin‐2 binding sites were not affected. In the frontal and parietal cortex, a significant correlation occurred between the latency to find the invisible platform in the water maze and somatostatin mRNA (r= ‐0.54 and 0.59 respectively, P > 0.02). These results indicate that ageing rats with memory impairments display some of the features of the somatostatinergic deficits observed in Alzheimer's disease.
Consequences of Transient Focal Cerebral Ischaemia for Second Messenger and Neurotransmitter Binding in the Rat: Quantitative Autoradiographic Analysis of Forskolin, Dopamine D1 Receptor Binding and Cerebral Blood Flow Changes
Authors: Gartshore Gail , Dawson Deborah , Patterson James , Macrae I. Mhairi
Published: April 2006

In order to study the consequences of reperfusion for ischaemic brain injury, quantitative ligand binding autoradiography was carried out in a model of reversible focal cerebral ischaemia. Endothelin‐1 applied to the abluminal surface of the middle cerebral artery in anaesthetized Sprague‐Dawley rats induced severe focal ischaemia and subsequent reperfusion (assessed by blood flow tracers [99mTc]HMPAO and [14C]iodoantipyrine respectively) by 2 h after insult. Ligand binding autoradiography on consecutive sections demonstrated these blood flow changes to be associated with a significant reduction in forskolin binding throughout the middle cerebral artery territory (e.g. 25% in parietal cortex, 11% in dorsolateral caudate nucleus). The most marked losses in forskolin binding were in areas where ischaemia was severe and reperfusion was poor. However, the same changes in cerebral blood flow had no significant effect on D1 dopamine receptor binding (e.g. >2% reduction in the caudate nucleus). These data demonstrate that ligand binding characteristics are significantly affected as early as 2 h after insult, with evidence of differential sensitivity for forskolin and D1 dopamine binding. With regard to the consequences of reperfusion, comparison with our previous study of 2 h maintained ischaemia demonstrates reperfusion‐related salvage of dopamine and forskolin binding in the caudate nucleus but possible exacerbation of forskolin binding loss in the cortex.
Distribution of Components of the SNARE Complex in Relation to Transmitter Release Sites at the Frog Neuromuscular Junction
Authors: Boudier Jeanne‐Andrée , Charvin Nathalie , Boudier Jean‐Louis , Fathallah Mohammed , Tagaya Mitsuo , Takahashi Masami , Seagar Michael J.
Published: April 2006

At the frog neuromuscular junction, neurotransmitter release sites are regularly spaced at 1 μm intervals along the nerve terminal, directly facing postsynaptic folds which contain a high density of acetylcholine receptors. Immunostaining and laser confocal scanning microscopy were used to compare the distribution of presynaptic proteins implicated in exocytosis with that of fluorescent α‐bungarotoxin. Syntaxin, synaptosome‐associated 25 kDa protein and calcium channels were located predominantly at release sites. Synaptobrevin (vesicle‐associated membrane protein) was distributed in the cytoplasm of the nerve terminal, presumably in the packets of microvesicles associated with each active zone. N‐Ethylmaleimide‐sensitive fusion protein (NSF) and soluble NSF attachment proteins (αβSNAP) displayed a diffuse distribution throughout the terminal cytoplasm and also colocalized in distinct concentrated zones adjacent to the presynaptic membrane.
Localization of Dopamine D2 Receptor in Rat Spinal Cord Identified with Immunocytochemistry and In Situ Hybridization
Authors: van Dijken Henk , Dijk Jan , Voorn Pieter , Holstege Jan C.
Published: April 2006

In the present study the distribution of dopamine D2 receptors in rat spinal cord was determined by means of immunocytochemistry using an anti‐peptide antibody, directed against the putative third intracellular loop of the D2 receptor and in situ hybridization (ISH) using a [35S]UTP labelled anti‐sense riboprobe. With the immunocytochemical technique, labelling was confined to neuronal cell bodies and their proximal dendrites. Strongest labelling was present in the parasympathetic area of the sacral cord and in two sexually dimorphic motor nuclei of the lumbosacral cord, the spinal nucleus of the bulbocavernosus and the dorsolateral nucleus. Moderately labelled cells were present in the intermediolateral cell column, the area around the central canal and lamina I of the dorsal horn. Weak labelling was present in the lateral spinal nucleus and laminae VII and VIII of the ventral horn. Except for the two sexually dimorphic motornuclei of the lumbosacral cord labelled motoneurons were not encountered. With the ISH technique radioactive labelling was present in many neurons, indicating that they contained D2 receptor mRNA. The distribution of these neurons was very similar to the distribution obtained with immunocytochemistry, but with ISH additional labelled cells were detected in laminae III and IV of the dorsal horn, which were never labelled with immunocytochemistry. The present study shows that the D2 receptor is expressed in specific areas of the rat spinal cord. This distribution provides anatomical support for the involvement of D2 receptors in modulating nociceptive transmission and autonomic control. Our data further indicate that D2 receptors are not directly involved in modulating motor functions with the exception, possibly, of some sexual motor functions.
Long‐term Depression of Horizontal Connections in Rat Motor Cortex
Authors: Hess Grzegorz , Donoghue John P.
Published: April 2006

The possibility for long‐term depression (LTD) of synaptic transmission in layer 11/11 I horizontal connections within motor cortex was investigated using field potentials and intracellular recordings in rat brain slices. The LTD was induced by low‐frequency stimulation at 2 Hz for 10 min in sites displaced horizontally by 0.5 mm from the stimulating electrode. Response amplitude measured 25‐30 min after 2 Hz stimulation ended was 79% of baseline values (n= 13) at half maximal stimulation and 59% when 2 Hz stimulus intensity was doubled (n= 10). In 13/15 tested cases LTD in horizontal connections was specific to the activated pathway. Intracellular recordings from six neurons confirmed synaptic character of response depression. Horizontal connections in which LTD was induced retained the capability of increasing synaptic strength. Long‐term potentiation could be induced in previously depressed pathways by simultaneous theta burst stimulation of two converging horizontal inputs combined with transient local application of GABAA receptor antagonist bicuculline methiodide (mean increase: 45 ± 8%, n = 6) or by simultaneous theta burst stimulation of converging horizontal and vertical inputs (mean change: 26 5 6%, n = 5). These data demonstrate that activity‐dependent mechanisms may regulate bidirectionally the effectiveness of horizontal synaptic coupling between cortical neurons, thus forming a potential mechanism for plasticity of cortical connections and the representation patterns they support.
Hydroxyindole‐O‐methyltransferase in the Chicken Retina: Immunocytochemical‐ Localization and Daily Rhythm of mRNA
Authors: Guerlotté Jérôme , Grève Pierre , Bernard Marianne , Grechez‐Cassiau Aline , Morin Fabrice , Collin Jean‐Pierre , Voisin Pierre
Published: April 2006

In the vertebrate retina and pineal gland, melatonin production displays diurnal variations with high levels at night. Hydroxyindole‐O‐methyltransferase (HIOMT, EC 2.1.1.4) catalyses the last step of melatonin biosynthesis. In the present study, a cDNA encoding chicken HIOMT was used to examine the effects of environmental lighting on HIOMT mRNA expression in the chicken retina. A day/night rhythm of HIOMT mRNA level was observed, with an average 5‐fold increase during the night. Light strongly suppressed the night‐time rise in HIOMT mRNA concentration while darkness prevented its daytime fall. An antibody directed against chicken HIOMT was used for immunocytochemical identification of retinal melatoninergic cells. HIOMT immunoreactivity could be observed in rods as well as in cones. However, the lowest levels of HIOMT immunoreactivity were always observed in the accessory cones of double cones. A few HIOMT‐positive cell bodies could also be observed in the inner nuclear layer. Altogether, these data indicate that HIOMT gene expression in the retina is organized on a daily basis as a direct response to light, and that the different types of photoreceptors may not be equally involved in melatonin production.
Regulation of Intracellular Ca2+ in Response to Muscarinic and Glutamate Receptor Agonists During the Differentiation of NTERA2 Human Embryonal Carcinoma Cells into Neurons
Authors: Squires P. E. , Wakeman J. A. , Chapman H. , Kumpf S. , Fidock M. D. , Andrews P. W. , Dunne M. J.
Published: April 2006

Single cell microfluorimetry was used to study intracellular calcium ion signals ([Ca2+]i) evoked by acetylcholine (ACh), glutamate receptor agonists and by KCI‐induced membrane depolarization, during neuronal differentiation of the human embryonal carcinoma (EC) cell line, NTERA2. In undifferentiated NTERA2 EC cells, [Ca2+]i) was elevated in response to ACh, but not to the glutamate receptor agonists NMDA, kainate or AMPA. The ACh‐induced rise in [Ca2+]i) was dependent upon both Ca2+ influx and Ca2+ mobilization from cytoplasmic calcium stores. Three other human EC cell lines responded similarly to ACh but not to glutamate or KCI‐induced depolarization. In neurons derived from NTERA2 cells by retinoic acid induction, [Ca2+]i) signals were evoked by ACh, NMDA, kainate and by an elevation of the extracellular KCI concentration. As in undifferentiated EC cells, the ACh‐mediated increases in [Ca2+li were governed by both Ca2+ influx and Ca2+ mobilization. In contrast, the effects of NMDA, kainate and KCI did not involve intracellular Ca2+ mobilization. The appearance of glutamate and KCI responsiveness was not detected in non‐neuronal differentiated derivatives of NTERA2 cells. Using a number of pharmacologically defined muscarinic receptor antagonists we found that NTERA2 EC cells express M1, M3, M4 and possibly M5 receptor subtypes linked to changes in [Ca2+]i), whilst only M3 and M5 are present in NTERA2‐derived neurons. The results were supported by PCR analysis of the muscarinic mRNA species expressed in the cells. The data demonstrate that differentiation of NTERA2 EC cells into neurons involves the induction of functional glutamate receptors coupled to rises in [Ca2+]i), and changes in the expression of muscarinic ACh receptor subtypes.
Localization of Glial Cell Line‐derived Neurotrophic Factor (GDNF) mRNA in Embryonic Rat by In Situ Hybridization
Authors: Suvanto Petro , Hiltunen Jukka O. , Arumäe Umas , Moshnyakov Maxim , Sariola Hannu , Sainio Kirsi , Saarrna Mart
Published: April 2006

The localization of glial cell line‐derived neurotrophic factor (GDNF) mRNA was studied by in situhybridization in rat from embryonic (E) day E10 to E15. At E10, GDNF mRNA is found in the urogenital field and the cranial part of the gut. At E11, the most abundant expression of GDNF mRNA is seen in the epithelial cells of the second, third and fourth pharyngeal pouches, the third and fourth pharyngeal arches and pharynx. Also mesenchymal cells of the gut and mesonephric tubules contain GDNF mRNA. At E13, expression is observed in the mesenchymal cell layers of the oesophagus, intestine and stomach, the mesenchymal cells around the condensing cartilages and metanephric kidney mesenchyme. Also, the epithelia of Rathke's pouch and pharynx are intensely labelled. High expression of GDNF mRNA continues at El5 in kidney, gastrointestinal tract and cartilage. At that stage, GDNF mRNA is seen also in whisker pad and skeletal muscles. The distribution of GDNF mRNA in embryonic rat suggests important roles for GDNF in the early differentiation of the kidney tubules, the innervation of the gastrointestinal tract and the differentiation process of the cartilage and muscle. Our results indicate novel functions for GDNF outside the nervous system.
The Plasma Membrane Protein SNAP‐25, but not Syntaxin, is Present at Photoreceptor and Bipolar Cell Synapses in the Rat Retina
Authors: Brandstätter J. H. , Wässle H. , Betz H. , Morgans C. W.
Published: April 2006

The two neuronal plasma membrane proteins synaptosomal associated protein 25 kDa (SNAP‐25) and syntaxin form, in current models of synaptic vesicle exocytosis, a docking complex on the presynaptic plasma membrane. This docking complex interacts with the integral synaptic vesicle protein, synaptobrevin. We have examined, in the rat retina, the distribution of SNAP‐25 and syntaxin using light and electron microscopic immunocytochemistry. SNAP‐25 immunoreactivity was present in the outer and inner nuclear layers at the membranes of photoreceptor and amacrine cell somata, in the outer plexiform layer in the terminals of photoreceptor cells and throughout the inner plexiform layer in the terminals of bipolar and amacrine cells. In contrast, syntaxin immunoreactivity was not found in the terminals of photoreceptor and bipolar cells, but only in the terminals of amacrine cells. Because of the absence of detectable syntaxin from the terminals of photoreceptor and bipolar cells, we propose that either a novel syntaxin isoform or a syntaxin–like protein exists at the ribbon synapses formed by these neurons.
Reserpine Affects Differentially the Density of the Vesicular Monoamine Transporter and Dihydrotetrabenazine Binding Sites
Authors: Naudon L. , Raisman‐Vozari R. , Edwards R. H. , Leroux‐Nicollet I. , Peter D. , Liu Y. , Costentin J.
Published: April 2006

We have studied the effect of a single injection of reserpine (5 mg/kg, s.c.) on the synaptic vesicle monoamine transporter (VMAT) density in the rat striatum, using two labelling procedures: radioimmunolabelling with an antibody against VMAT, and binding of the specific ligand [3H]dihydrotetrabenazine ([3H]TBZOH). In the rostral and medial striatum, the distribution of VMAT immunoreactivity displayed the highest density in the lateral subregions. In the caudal part of the striatum, VMAT immunoreactivity showed increasing density from dorsal to ventral subregions. The VMAT immunoreactivity was not altered 2 and 30 days after the reserpine injection, whereas [3H]TBZOH binding site density, measured on adjacent slices, showed a dramatic decrease at day 2 and a moderate recovery at day 30, suggesting that despite a persistent blockade of [3H]TBZOH binding sites, VMAT protein density was unchanged.
The Effects of Neonatal Dorsal Root Section on the Survival and Dendritic Development of Lumbar Motoneurons in the Rat
Authors: O'Hanlon G. M. , Lowrie M. B.
Published: April 2006

Peripheral nerve crush during the early neonatal period results in the death of a large proportion of affected motoneurons and abnormal dendritic development in those which survive. The present study reports the effects of neonatal dorsal root section on motoneurons supplying the extensor digitorum longus muscle of the rat. This lesion did not result in motoneuron death, but did disrupt subsequent dendritic development. In cells retrogradely labelled with cholera toxin subunit B conjugated to horseradish peroxidase, there was little change in adult dendritic morphology in the transverse plane, where abnormalities associated with loss of efferent contact and cell death have been found. However, there was a caudal expansion of the dendritic field, an effect seen following nerve crush but not after blockade of neuromuscular transmission alone. The results show that disruption of dorsal root sensory inputs alone can affect the dendritic development of motoneurons but does not cause their death. In conjunction with our earlier findings, it is clear that both afferent and efferent connections are required for normal dendritic development, and disruption of either has a characteristic effect on survival and dendritic morphology.
Neurite Outgrowth on Non‐permissive Substrates In Vitro is Enhanced by Ectopic Expression of the Neural Adhesion Molecule L1 by Mouse Astrocytes
Authors: Mohajeri M. Hasan , Bartsch Udo , van der Putten Herman , Sansig Gilles , Mucke Lennart , Schachner Melitta
Published: April 2006

Axonal regrowth in the lesioned central nervous system (CNS) of adult mammals is, in part, prevented by non‐permissive properties of glial cells and myelin. To test if ectopic expression of the neurite outgrowth promoting recognition molecule L1 will overcome these non‐permissive influences and promote neurite outgrowth, L1 was expressed in astrocytes of transgenic mice using regulatory sequences of the glial fibrillary acidic protein (GFAP) gene. Northern blot analysis of different transgenic lines revealed different levels of transgenically expressed L1. Cultured astrocytes derived from transgenic animals displayed L1 immunoreactivity at the cell surface and in situ hybridization and immunocytochemical analysis of optic nerves from adult transgenic mice localized L1 expression to astrocytes. Expression of L1 protein by transgenic astrocytes was significantly upregulated in lesioned optic nerves. When mouse small cerebellar neurons or chick dorsal root ganglion neurons were cultured on cryosections of lesioned optic nerves or astrocyte monolayers from transgenic mice, respectively, neurite outgrowth was increased up to 400% on tissue sections and 50% on astrocytes compared with similar preparations from non‐transgenic mice. The increase in neurite outgrowth on tissue sections or astrocyte monolayers from different transgenic lines was proportional to the different levels of L1 expression. Moreover, increased neurite outgrowth on these substrates was specifically inhibited by polyclonal L1 antibodies. In vivo, rescue of severed axons was enhanced in transgenic versus wild type animals, while regrowth of axons was slightly, but not significantly, increased. Together, our observations demonstrate that L1 promotes neurite outgrowth when expressed ectopically by astrocytes and that L1 is able to overcome, at least partially, the non‐permissive substrate properties of differentiated CNS glial cells in vitro.