bluuebunny/crossref_metadata_2025 huggingface dataset:
Alpha1‐adrenergic Effects on Dopamine Neurons Recorded Intracellularly in the Rat Midbrain Slice
Authors: Grenhoff Johan , North R. Alan , Johnson Steven W.
Published: April 2006

Previous studies have indicated excitatory adrenergic effects on midbrain dopamine systems. To investigate the cellular mechanisms, intracellular recordings were made from neurons in perfused, oxygenated slices of male rat midbrain. Electrophysiological and pharmacological parameters were used to identify cells as principal (presumably dopaminergic) neurons as opposed to secondary (presumably GABAergic) neurons in the substantia nigra zona compacta and the ventral tegmental area. Noradrenalin (10–100 μM) hyperpolarized 57% of all principal cells and depolarized 36%. Sulpiride (100–1000 nM), a dopamine D2 receptor antagonist, completely blocked noradrenalin‐induced hyperpolarizations (six of six cells). In sulpiride, noradrenalin depolarized 58% of all principal neurons and had no effect in 42%; this effect was mimicked by the α‐adrenergic agonist phenylephrine (10–30 μM) which depolarized 43 of 72 cells. The α1 receptor antagonist prazosin (30–100 nM) completely blocked the membrane depolarization produced by either noradrenalin or phenylephrine in all cells tested, whereas α2‐ and β‐adrenergic agents had no effect. In voltage clamp, phenylephrine evoked an inward current (at ‐60 mV) and reduced cord conductance by 0.81 ± 0.14 nS (n= 4). Inward current evoked by phenylephrine became outward at ‐96 ± 8 mV, which is near the membrane reversal potential for potassium as predicted by the Nernst equation. Phenylephrine also depolarized secondary cells and increased the frequency of spontaneous GABAA receptor‐mediated postsynaptic potentials recorded in both principal and secondary cells. We conclude that stimulation of α1‐adrenergic receptors depolarizes principal (dopamine) neurons by reducing membrane conductance for potassium, but this effect is modulated by the increase in frequency of spontaneous inhibitory postsynaptic potentials evoked by stimulation of α1‐adrenergic receptors located on local interneurons.
Development of Electrical Membrane Properties and Discharge Characteristics of Superior Olivary Complex Neurons in Fetal and Postnatal Rats
Authors: Kandler Karl , Friauf Eckhard
Published: April 2006

Although hearing onset occurs relatively late during ontogeny of rats [around postnatal day (P) 12], anatomical brainstem connections are formed much earlier and are present before birth, indicating that a substantial amount of maturation occurs without acoustic input. Electrical activity is thought to influence neuronal development, but the physiological properties of auditory brainstem neurons during perinatal maturation are barely known. The present study focuses on the development of electrophysiological membrane properties of neurons in the rat's superior olivary complex (SOC), the first binaural station in the mammalian auditory brainstem. In in vitro slice preparations, intracellular recordings were obtained from 115 SOC cells from embryonic day (E) 18 to P17, and cells were morphologically identified by intracellular injection of biocytin or neurobiotin. By E18, i.e. 4 days before birth, SOC neurons were capable of generating Na+‐dependent action potentials. Several passive and active membrane properties, including the resting potential, spike threshold and spike amplitude, did not change with development. In contrast, input resistance, time constant and spike duration decreased significantly, and maximal spike frequency increased significantly during the age period sampled. Our results show that rat SOC neurons display mature as well as immature electrical membrane properties during the same developmental period when anatomical connections are refined and when the soma‐dendritic morphology develops. We conclude, therefore, that their membrane properties represent adequate physiological adaptations to the immature auditory brainstem microcircuits and that they form a basis upon which the development of these microcircuits is shaped.
Developmental Regulation of Acetylcholinesterase Transcripts in the Mouse Diaphragm: Alternative Splicing and Focalization
Authors: Legay Claire , Huchet Monique , Massoulié Jean , Changeux Jean‐Pierre
Published: April 2006

We studied the splicing and compartmentalization of acetylcholinesterase (AChE) mRNAs during muscle differentiation in the mouse, both in vitro and in vivo. We used the polymerase chain reaction (PCR) to analyse AChE mRNAs in cultures of the myogenic C2 and So l8 cell lines, and in the developing diaphragm, from embryonic day 14 (E14). We characterized three types of alternatively spliced AChE mRNAs, encoding catalytic subunits that differ by their C‐terminal regions (R, H and T). The T transcript is predominant in all cases and represents the only AChE mRNA in the adult muscle. We detected the presence of the minor R and H transcripts in the myogenic cell lines, both as myoblasts and differentiated myotubes, and also in the diaphragm from E14 until birth. At E14 the R transcript represents ∼1% of AChE mRNA and the level of the H transcript is still lower. By in situ hybridization, we found that the T AChE mRNAs begin to preferentially accumulate at the level of the first neuromuscular contacts in the mouse diaphragm and other muscles as early as E14, e.g. concomitantly with mRNAs encoding the receptor subunits. This suggests that a common control mechanism ensures the synaptic focalization of mRNAs encoding the cholinergic proteins AChE and acetylcholine receptor during muscle development.
NADPH Diaphorase‐Containing Nonpyramidal Cells in the Rat Hippocampus Exhibit Differential Sensitivity to Kainic Acid
Authors: Miettinen R. , Kotti T. , Halonen T. , Riekkinen P.
Published: April 2006

Neurons containing nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase exhibit high resistance to several excitotoxins. In the neocortex and striatum, however, these neurons are sensitive to kainic acid. Here we report that, 2 weeks after i.p. injection of kainic acid, the number of NADPH diaphorase neurons in the hilus and CA1 subfield was decreased, whereas the cell counts in the other hippocampal areas were to a great extent similar to those for the controls. We propose that the loss of NADPH diaphorase neurons in the hippocampus after systemic injection of kainic acid is associated with the pathophysiological processes involved in the spreading of epileptic seizure activity rather than to the direct neurotoxic effect of the kainic acid per se.
Cerebral Energy Metabolism and Immediate Early Gene Induction Following Severe Incomplete Ischaemia in Transgenic Mice Overexpressing the Human Ornithine Decarboxylase Gene: Evidence that Putrescine is not Neurotoxic In Vivo
Authors: Lukkarainen Jouko , Kauppinen Risto A. , Koistinaho Jari , Halmekytö Maria , Alhonen Leena , Jänne Juhani
Published: April 2006

Cerebral ischaemia causes activation of ornithine decarboxylase followed by accumulation of putrescine, and these biochemical phenomena have been thought to contribute to the development of neuronal damage. We have used a transgenic mouse line overexpressing the human ornithine decarboxylase gene in their neurons with constitutively high putrescine to study the possible role of putrescine in development of neuronal damage in forebrain ischaemia. An incomplete forebrain ischaemia model was developed in which common carotid arteries were bilaterally occluded and reduction of blood pressure caused by orthostatic reaction was used as a way of decreasing cerebral circulation. Cerebral high‐energy metabolites, intracellular pH and lactate were monitored by means of 31P and 1H nuclear magnetic resonance spectroscopy respectively. Incomplete ischaemia for 15 min resulted in severe energy failure, as indicated by an increase in the inorganic phosphate/phosphocreatine ratio, intracellular acidification from a pH of ∼7.1 to ∼6.5 and an increase in lactate concentration from <1 to ∼10 mmol/kg in both syngenic and transgenic mice. Following deocclusion, recovery of energy metabolites, intracellular pH and lactate were identical in both animal groups. Ornithine decarboxylase activity rose 9‐ and 3‐fold in syngenic and transgenic mice respectively 6 h after ischaemia. Activation of ornithine decarboxylase resulted in extensive accumulation of putrescine in the brains of transgenic animals 12–24 h after ischaemia, which was ∼50‐fold greater than the basal level in syngenic mice. In situ hybridization experiments revealed induction of transcription factors c‐Fos and zif‐268 in the hippocampus, throughout the cerebral cortex and striatum 1–3 h after ischaemia. Messenger RNA of heat shock protein 70 was induced in dentate gyrus and CA3 and CA4 subfields of the hippocampus 1 h after ischaemia. The distribution and extent of induced mRNAs were similar in syngenic and transgenic animals. Histological evaluation did not reveal any difference between the two animal groups despite large variation in their cerebral putrescine content. Neuronal necroses were observed in the CA4 layer of hippocampi in both syngenic and transgenic mice. These data suggest that ornithine decarboxylase activation and accumulation of endogenous putrescine are indicative of an ischaemic insult and that these changes reflect an adaptive response rather than acting as causative factors of neuronal damage.
Neonatal Rat Cerebellar Granule and Purkinje Neurons in Culture Express Different GABAA Receptors
Authors: Zempel John Martin , Steinbach Joe Henry
Published: April 2006

We have established a culture system for microexplants of rat cerebellar cortical tissue in which cells develop morphologically, express type‐A receptors for the inhibitory neurotransmitter γ‐aminobutyric acid (GABA) and form GABAergic synaptic connections. Criteria of cell size and shape allow reliable identification of granule and Purkinje neurons, criteria confirmed by studies of the binding of antibodies to calbindin D28K and GABA. Both granule and Purkinje neurons express GABAA receptors, but granule neurons fall into two classes in terms of their sensitivity. Granule neurons which do not show spontaneous synaptic currents are relatively insensitive to GABA, while granule neurons with synaptic currents are much more sensitive. The responses of Purkinje neurons to applications of 1 μM GABA are relatively insensitive to Zn2+ ions (10 μM), and are potentiated by chlordiazepoxide (100 μM) and La3+ ions (100 μM). Responses of innervated granule neurons, on the other hand, are blocked more strongly by Zn2+ ions, are less affected by chlordiazepoxide and are equally potentiated by La3+ ions. Hence these cultures provide a source of identifiable, functionally innervated cells which express distinct types of GABAA receptors.
Implication of a Nitric Oxide Synthase Mechanism in the Action of Substance P: L‐NAME Blocks Thermal Hyperalgesia Induced by Endogenous and Exogenous Substance P in the Rat
Authors: Radhakrishnan V. , Yashpal K. , Hui‐Chan C. W. Y. , Henry J. L.
Published: April 2006

The effects of i.p. administration of the nitric oxide synthase inhibitor NG‐nitro‐L‐arginine methylester (L‐NAME) and its inactive isomer, D‐NAME, were tested in two nociceptive paradigms in the rat. In the first paradigm, rats were lightly anaesthetized with a mixture of chloral hydrate (120 mg/kg, i.p.) and sodium pentobarbital (20 mg/ kg, i.p.). Tail flick reaction times were monitored and thermal hyperalgesia was induced by immersion of the tail in hot water at 55°C for 1.5 min. In the groups of rats pretreated with saline (n= 5), 100 mg/kg D‐NAME (n= 6), 10 (n= 5) or 25 (n= 6) mg/kg L‐NAME, this thermal injury induced a transient reduction in the reaction time that was 54–59% of the baseline value. However, in the groups of rats pretreated with 50 (n= 6) or 100 (n= 7) mg/kg L‐NAME the reaction times were 73.9 ± 2.7% (P < 0.05) and 102.3 ± 0.9% (P < 0.001) of the baseline values respectively, indicating a block of the hyperalgesic responses seen in the other groups. As this hyperalgesia has been reported to be blocked by NK‐1 receptor antagonists, it is suggested that it is due to the action of endogenous substance P. In the second paradigm, tail flick responses were monitored in the awake rat and thermal hyperalgesia was induced by intrathecal administration of substance P (6.5 nmol) via a chronically implanted catheter. In the group of rats pretreated with saline (n= 5) or D‐NAME (n= 5; 100 mg/kg), substance P reduced the reaction time to 39.1 ± 9.9 and 45.5 ± 2.1% of the baseline value respectively. However, in the rats pretreated with L‐NAME (n= 6; 100 mg/kg), the reaction time following substance P administration was 108.8 ± 8.8% of the baseline value (P < 0.001), indicating a block of the hyperalgesic response induced by substance P. These data indicate that thermal hyperalgesia induced by endogenously released or exogenously administered substance P, are blocked by L‐NAME but not by its enantiomer, D‐NAME. Therefore an involvement of a nitric oxide synthase mechanism in the hyperalgesic responses to substance P is suggested.
Miniature Endplate Current Kinetics at the Mouse Neuromuscular Junction: Effects of Temperature and Medium Viscosity
Authors: Tanzi Franco , D'Angelo Egidio
Published: April 2006

The time course of miniature endplate currents (MEPCs), derived by extracellular focal recording, was studied in the mouse neuromuscular junction at different temperatures and medium viscosities, and in eserine‐treated endplates. At low temperatures (6–10°C), almost the whole MEPC decay is exponential and the rising phase is not significantly modified by the channel closing process. At physiological temperatures (38–40°C), the early part of the decay is much slower than the later part and the rising phase is made shorter and smaller by channel closing, showing that the channel opening and channel closing processes overlap remarkably. Even at physiological temperatures, however, the late part of MEPC decay shows an exponential time course. At high temperatures the channel opening process has low temperature sensitivity and slows down when bath solution viscosity is increased, suggesting that at high temperatures channel opening kinetics may mainly be controlled by the time course of acetylcholine diffusion. The lower limit of conformational change rate leading to channel opening was estimated at 10°C (4400 s‐1). Experimentally recorded MEPCs were mathematically simulated to obtain a quantitative description of the processes controlling MEPC generation. Mathematical simulation further suggests that (i) acetylcholine diffusion kinetics may affect the onset rate of MEPCs without, however, being rate‐limiting; and (ii) partial, transient acetylcholinesterase inhibition operated by acetylcholine may explain the low temperature sensitivity exhibited by the onset rate of MEPCs at high temperatures.
Somatosensory Activations of the Parietal Operculum of Man. A PET Study
Authors: Ledberg Anders , O'Sullivan Brendan T. , Kinomura Shigeo , Roland Per E.
Published: April 2006

We tested the hypothesis that somatosensory discrimination of roughness (microgeometry) but not of shape (macrogeometry) would activate the parietal operculum (PO) in man. It was also investigated whether a simple square pulse indentation of the skin on the index finger would activate the PO. Regional cerebral blood flow was measured with [15O]butanol and positron emission tomography in a total of 20 normal volunteers. Ten subjects used their right hand to discriminate objects that differed in roughness and similar smooth objects that differed in length. Ten other subjects pressed a button when they felt a square pulse indentation of the skin on their right index finger in a somatosensory reaction time task. Discrimination of roughness activated one field in the PO contralaterally and two fields ipsilaterally to the stimulated hand. The discrimination of length activated one field in the PO located ipsilaterally to the stimulated hand. The somatosensory reaction time task also activated one contralateral and two ipsilateral fields in the PO, and these fields partially overlapped the activated fields in the roughness discrimination task. Based on the extension of these fields and their overlaps we conclude that there exist at least one part of the contralateral PO and at least two parts of the ipsilateral PO that can be activated by somatosensory stimulation of the right hand. We argue further that the contralateral activated part contains a region that can be activated by roughness.
In Vivo Modulation of Sigma Receptor Sites by Calcitonin Gene‐related Peptide in the Mouse and Rat Hippocampal Formation: Radioligand Binding and Electrophysiological Studies
Authors: Bouchard Pascale , Monnet François , Bergeron Richard , Roman François , Junien Jean‐Louis , de Montigny Claude , Debonnel Guy , Quirion Rémi
Published: April 2006

Possible interactions between sigma (σ) receptor sites and calcitonin gene‐related peptides (CGRP) were investigated using receptor subtype‐related analogues and fragments in in vivo [3H](+)SKF 10 047/σ binding in the hippocampus, and electrophysiological recording of the N‐methyl‐D‐aspartate (NMDA)‐induced activation of CA3 pyramidal neurons, two well‐established σ assays. In both paradigms, CGRP and the agonist [Cys(ACM)2,7]hCGRPα modulated σ systems. In vivo binding experiments demonstrated that CGRP and [Cys(ACM)2,7]hCGRPα inhibited 25–40% of specific [3H](+)SKF 10 047 labelling in the mouse hippocampal formation while the purported antagonist hCGRP8–37 was inactive. The specificity of this modulation was demonstrated further by the lack of effect of other vasoactive peptides, including the atrial natriuretic peptide, substance P, and its N‐terminal fragment, substance P1–7. In the CA3 subfield of the rat dorsal hippocampus, hCGRPα decreased (up to 61%) the NMDA‐induced activation of the pyramidal neurons. Conversely, the linear analogue [Cys(ACM)2,7]hCGRPα enhanced (by 85%) the NMDA‐induced activation of CA3 pyramidal neurons, while the antagonistic fragment hCGRP8–37 had no effect. Haloperidol, a high‐affinity σ receptor ligand, inhibited by 90% the in vivo [3H](+)SKF 10 047 labelling, and prevented the modulation of the NMDA‐induced activation by hCGRPα and [Cys(ACM)2,7]hCGRPα. It thus appears that CGRP can modulate σ‐related systems in the hippocampal formation.
Properties of unitary IPSPs evoked by anatomically identified basket cells in the rat hippocampus
Authors: Buhl E. H. , Cobb S. R. , Halasy K. , Somogyi P.
Published: April 2006

Hippocampal pyramidal cells receive GABA‐mediated synaptic input from several distinct interneurons. In order to define the effect of perisomatic synapses, intracellular recordings were made with biocytin‐containing microelectrodes from synaptically connected inhibitory and pyramidal cell pairs in subfields CA1 and CA3 of the rat hippocampus. Subsequent physiological analysis was restricted to the category of cells, here referred to as basket cells (n= 14), which had an efferent synaptic target profile (n= 282 synaptic contacts) of predominantly somatic (48.2%) and proximal dendritic synapses (45.0%). Electron microscopic analysis revealed that in two instances identified postsynaptic pyramidal cells received a total of 10 and 12 labelled basket cell synapses respectively. At an average membrane potential of ‐57.8 ± 4.6 mV, unitary inhibitory postsynaptic potentials (IPSPs; n= 24) had a mean amplitude of 450 ± 238 μV, a 10–90% rise time of 4.6 ± 3.2 ms and, measured at half‐amplitude, a mean duration of 31.6 ± 18.2 ms. In most instances (n= 19) the IPSP decay could be fitted with a single exponential with a mean time constant of 32.4 ± 18.0 ms. Unitary basket cell‐evoked IPSPs fluctuated widely in amplitude, ranging from the level of detectability to <2 mV. The response reversal of IPSPs (n= 5) was extrapolated to be at ‐74.9 ± 6.0 mV. Averages of unitary IPSPs had a mean calculated conductance of 0.95 ± 0.29 nS, ranging from 0.52 to 1.16 nS. Unitary basket cell IPSPs (n= 3) increased in amplitude by 26.3 ± 19.9% following bath application of the GABAB receptor antagonist CGP 35845A (1–4 μM), whereas subsequent addition of the GABAA receptor antagonist bicuculline (10–13 μM) reduced the IPSP amplitude to 13.5 ± 3.1% of the control response. Rapid presynaptic trains of basket cell action potentials resulted in the summation of up to four postsynaptic responses (n= 5). However, any increase in the rate of tonic firing (2‐ to 10‐fold) led to a <50% reduction of the postsynaptic response amplitude. At depolarized membrane potentials, averaged IPSPs could be followed by a distinct depolarizing overshoot or postinhibitory facilitation (n= 4). At firing threshold, pyramidal cells fired postinhibitory rebound‐like action potentials, the latter in close temporal overlap with the depolarizing overshoot. In conclusion, hippocampal basket cells have been identified as one source of fast, GABAA receptor‐evoked perisomatic inhibition. Unitary events are mediated by multiple synaptic release sites, thus providing an effective mechanism to avoid total transmission failures.
The Axonal Arborization of Single Thalamic Reticular Neurons in the Somatosensory Thalamus of the Rat
Authors: Pinault Didier , Bourassa Jacques , Deschênes Martin
Published: April 2006

This study describes the axonal projections of single neurons of the thalamic reticular complex within the somatosensory thalamic nuclei in rats. Experiments were performed under urethane anaesthesia and reticular cells were labelled by extracellular microiontophoretic applications of biocytin. The axonal arborization of 25 thalamic reticular cells projecting to the ventrobasal (VB) nucleus and/or to the posterior thalamic (PO) complex were reconstructed from serial horizontal sections. Reticular cells labelled with biocytin display somatodendritic features similar to those reported previously. Their cell body is fusiform and their dendrites bear few spines and show a high degree of streaming along the horizontal curved axis of the nucleus. In most cells, axon‐like beaded processes stem out from dendrites but, contrary to previous descriptions, no intrareticular axonal collateral was observed. The axonal arborization of most thalamic reticular cells is confined within the limits of a single thalamic nucleus; only two neurons were seen projecting to both the VB and the Po nuclei. In VB, termination fields form short rods (diameter ∼ 150 μm, length ∼ 200‐300 μm) densely packed with grape‐like boutons and varicosities; termination fields in Po are larger, much less dense, and they are contained within a horizontal slab of tissue (thickness ∼ 200 μm, mediolateral width ∼ 400 μm, rostrocaudal length ∼ 1 mm. By charting the position of all labelled cells within the thickness of the thalamic reticular complex, a strip‐like arrangement was revealed. Cells projecting to Po occupy the innermost portion of the nucleus whereas those projecting to the ventral‐posteromedial and ventral‐posterolateral nuclei are located respectively in the middle and in the outer tiers of the nucleus. This strip‐like reciprocity was confirmed by separate biocytin injections performed in VB and in Po. These results show that inhibition of reticular origin is distributed within the rat dorsal thalamus in a highly specific manner, most likely according to a principle of reciprocity within the somatotopic representation of the body.
Associative Synaptic Potentials in the Piriform Cortex of the Isolated Guinea‐pig Brain In Vitro
Authors: Biella Gerardo , de Curtis Marco
Published: April 2006

The involvement of local and remote associative fibres in the generation of piriform cortex synaptic potentials was investigated in the isolated guinea‐pig brain maintained in vitro by arterial perfusion by implementing current source density analysis (CSD) on cortical field potential profiles. Previous hypotheses were verified using acute surgical isolation of piriform cortical areas to study different synaptic events separately. Stimulation of the lateral olfactory tract activated associative potentials throughout the piriform cortex. In the anterior piriform cortex, the current sinks responsible for the generation of associative potentials were located in the superficial portion of layer lb and in layer III. In the posterior piriform cortex, two associative events were observed: an early sink located in the superficial part of layer Ib, followed by a sink in the deep part of the same layer. In the anterior piriform cortex, local associative synaptic potentials were separated from the component carried by long projective fibres by surgically isolating a small area of cortex monosynaptically activated by lateral olfactory tract stimulation. In this patch of lateral olfactory tract‐connected anterior piriform cortex, local associative sinks were observed in the superficial lb layer and in layer III. Monosynaptic activation of the isolated patch of anterior piriform cortex induced purely associative potentials throughout the piriform cortex. These potentials were mediated by the synaptic activation of apical dendrites in the superficial lb layer and selectively abolished by severing the long associative fibres. The anterior piriform cortex layer III sink and the posterior piriform cortex deep lb associative component were evoked by the activation of large population spikes in the monosynaptic anterior piriform cortex and the disynaptic posterior piriform cortex response respectively. These two sinks are presumably generated locally through a polysynaptic circuit, whose activation depends on the degree of cortical excitation. Olfactory signal processing in the guinea‐pig piriform cortex during states of normal excitability is supported by the interactions between associative inputs impinging on the synapses located separately on the dendrites of pyramidal neurons. An increase in the synchronization of piriform cortex neuron discharge activates usually silent local circuit synapses.
Effect of Collicular Proteoglycan on the Survival of Adult Rat Retinal Ganglion Cells Following Axotomy
Authors: Huxlin Krystel R. , Dreher B. , Schulz M. , Sefton A. Jervie , Bennett M. R.
Published: April 2006

Consistent with numerous previous studies, we have found that in adult rats 29% of cells retrogradely prelabelled by injections into retino‐recipient nuclei are lost 1 week after intraorbital section of the optic nerve. This figure increases to 76% 2 weeks after axotomy. Intraocular injections of 150 ng of 480 kda chondroitin sulphate proteoglycan purified from the superior colliculi of neonatal rats were performed every third day after axotomy. This procedure resulted in the loss of only 3 and 28% of the axotomized retinal ganglion cells 7 and 14 days respectively after optic nerve section. Intraocular injections of chondroitin sulphate type C, one of the sugar types present on the collicular proteoglycan, also resulted in a significant saving of axotomized ganglion cells (with the loss of only 48% 14 days after optic nerve lesion). These findings suggest that the collicular proteoglycan, and to a lesser extent its sugar moieties, substantially slows down the degeneration of adult retinal ganglion cells following axotomy.
Quantitative Comparison of the Transient Rescue Effects of Neurotrophic Factors on Axotomized Motoneurons In Vivo
Authors: Vejsada R. , Sagot Y. , Kato A. C.
Published: April 2006

A reproducible neuronal degeneration induced by nerve lesion in neonatal rats or mice provides a convenient in vivo assay for testing the survival‐promoting activity of putative growth factors on motoneurons. The goal of this study was to compare the rescue effects of the four known neurotrophins [nerve growth factor (NGF), brain‐derived neurotrophic factor (BDNF), neurotrophin‐3 (NT‐3) and neurotrophin‐4 (NT‐4)] and two of the cytokines [ciliary neurotrophic factor (CNTF) and leukaemia inhibitory factor (LIF)] in one particular experimental model of spinal motoneuron degeneration at two different survival times. The sciatic nerve was cut in neonatal rats and the factors were applied onto the nerve stump; bovine serum albumin was used in controls. Simultaneous application of the retrograde tracer fluoro‐gold made it possible to count motoneurons specifically in the sciatic pool. One week after lesion, the neurotrophins BDNF, NT‐3 and NT‐4, but not NGF, equally enhanced motoneuron survival compared to controls; their effects were significantly better than those of the cytokines. However, the rescue from cell death was only transitory because a great number of the motoneurons died during the second week after nerve lesion. Additional BDNF and/or CNTF supplied by repeated subcutaneous injections (1 mg/ml) over 2 weeks could not prevent this delayed motoneuron loss. These results suggest that still other factors or alternative routes of administration may be required for permanent rescue of the lesioned immature motoneurons.
NGF Deprivation of Adult Rat Brain Results in Cholinergic Hypofunction and Selective Impairments in Spatial Learning
Authors: Van der Zee Catharina E. E. M. , Lourenssen Sandra , Stanisz Jolanta , Diamond Jack
Published: April 2006

Cholinergic hypofunction has often been correlated with a variety of behavioural impairments. In the present study, adult Wistar rats were intraventricularly infused with antibodies to nerve growth factor (anti‐NGF) to examine the effects on cholinergic neurons of the basal forebrain, and on behavioural performance. Immunocytochemical techniques indicated that chronically infused anti‐NGF penetrates into the basal forebrain, cortex, striatum, corpus callosum and hippocampus, confirming previous findings after a single injection. Treatment with anti‐NGF for 1 or 2 weeks resulted in a significant decrease of 27‐33% in density of choline acetyltransferase immunostaining of the cholinergic cell bodies in the medial septum and vertical diagonal band, and a 26% reduction in choline acetyltransferase enzyme activity in the septal area. An array of spatial learning Morris water maze tasks was used to distinguish between acquisition skills and the flexible use of learned information in novel tests. Rats subjected to the spatial learning paradigm received anti‐NGF infusion for 2 weeks prior to and for another 2 weeks during the behavioural testing. The anti‐NGF‐treated animals were found to be no different from those receiving control serum in the Morris water maze acquisition task, either in the latency to find the platform or in the time spent searching in the training quadrant when the platform was removed. However, in consecutive extinction trials, anti‐NGF rats continued to search in the empty training quadrant, suggesting the occurrence of perseveration; control rats expanded their search over other areas of the pool. This inflexibility of the anti‐NGF rats was also evident from their difficulty in learning to find a relocated platform in the reversal task. Finally, the anti‐NGF‐treated animals demonstrated hyperactivity in the open field, resembling in this respect the behaviour of animals after septal and fimbria fornix lesions, and during pharmacological cholinergic blockade. While these data support a role for NGF in cholinergic function and spatial learning behaviour, they indicate that deficits in the latter, e.g. those demonstrated in impaired aged rats, may not be attributable selectively to deficits in the function of NGF‐sensitive cholinergic neurons of the basal forebrain.
Exocytosis and Selective Neurite Calcium Responses in Rat Cerebellar Granule Cells During Field Stimulation
Authors: Cousin Michael A. , Held Brigitte , Nicholls David G.
Published: April 2006

The free calcium concentration, [Ca2+]c, in fura‐2‐loaded rat cerebellar granule cells was investigated by digital imaging during trains of uniform field stimuli in order to compare the ability of calcium channels in somata and neurites to respond to brief, physiologically relevant depolarizations. Very few somata responded to 20 Hz trains of 1 ms pulses, while virtually all neurites showed an extensive increase which was rapidly reversed when stimulation was terminated. In contrast, both somata and neurites responded when cells were depolarized with 50 mM KCl. The field stimuli evoked a tetrodotoxin‐sensitive increase in Na+ concentration in both somata and neurites. When 4‐aminopyridine, which inhibits delayed K+ currents in these cells, was present during the field stimulus both somata and neurites increased their [Ca2+]c, suggesting that prolongation of the duration of depolarization is required for somatic Ca2+ channel activation. The neurite response did not depend on the orientation of the neurite relative to the applied field. The neurite response was insensitive to nifedipine (1 μM) and ω‐agatoxin‐IVA (30 nM) but was uniformly inhibited by ω‐conotoxin‐GVIA (30% inhibition at 1 μM) and ω‐conotoxin‐MVIIC (44% inhibition at 5 μM). The two inhibitors were not additive. The neurite [Ca2+]c response was insensitive to the combination of ionotropic glutamate receptor antagonists. Field stimulation caused the exocytosis of the fluorescent probe FM1‐43 previously loaded during KCl depolarization, suggesting that presynaptic Ca2+ channels contribute to the field‐evoked neurite response.
Ammonia Prevents Activation of NMDA Receptors by Glutamate in Rat Cerebellar Neuronal Cultures
Authors: Marcaida Goizane , Miñana María‐Dolores , Burgal María , Grisolía Santiago , Felipo Vicente
Published: April 2006

Acute ammonia toxicity is mediated by activation of NMDA receptors and is prevented by chronic moderate hyperammonaemia. The aim of this work was to assess whether the protective effect of chronic hyperammonaemia is due to impaired activation of the NMDA receptor. It is shown that chronic hyperammonaemia in rats decreases the binding of [3H]MK‐801 to synaptosomal membranes from the hippocampus but not the amount of NMDAR1 receptor protein as determined by immunoblotting. In primary cultures of cerebellar neurons, long‐term treatment with 1 mM ammonia also decreased significantly the binding of [3H]MK‐801. These results suggest that ammonia impairs NMDA receptor activation. To confirm this possibility we tested the effect of long‐term treatment of the cultured neurons with 1 mM ammonia on three well known events evoked by activation of the NMDA receptor: neuronal death induced by glutamate, increase in aspartate aminotransferase activity and increase in free intracellular [Ca2+]. Long‐term treatment with ammonia prevented noticeably the effects of glutamate or NMDA on all these parameters. These results indicate that long‐term treatment of neurons with 1 mM ammonia leads to impaired function of the NMDA receptor, which cannot be activated by glutamate or NMDA. Activation of protein kinase C by a phorbol ester restored the ability of the NMDA receptor to be activated in neurons treated with ammonia. This suggests that ammonia impairs NMDA receptor function by decreasing protein kinase C‐dependent phosphorylation.
An NK1 Receptor‐dependent Component of the Slow Excitation Recorded Intracellularly from Rat Motoneurons Following Dorsal Root Stimulation
Authors: Baranauskas G. , Traversa U. , Rosati A. M. , Nistri A.
Published: April 2006

Intracellular recording from lumbar motoneurons of the neonatal rat spinal cord in vitro was used to study how recently developed non‐peptide antagonists such as SR‐140333 and SR‐48698, known to block distinct subtypes of tachykinin receptors peripherally, might affect synaptic transmission elicited by electrical stimulation of dorsal root fibres. SR‐140333 (1 μM) preferentially antagonized responses mediated by an exogenously applied agonist acting on the NK1 receptor subclass, while SR‐48968 (0.5 μM) preferentially reduced responses mediated by an exogenously applied agonist acting on the NK2 receptor subclass. SR‐48968 did not affect fast or slow excitatory postsynaptic potentials (EPSPs) or ‘wind‐up’responses induced by repetitive, low‐frequency stimulation (mimicking certain types of nociceptive input); binding studies using this radiolabelled ligand disclosed specific binding activity (21 fmol/mg protein) selectively displaced by an NK2 receptor agonist. SR‐140333 reduced the late component of fast and slow EPSPs, and of wind‐up. Pharmacological block of ionotropic glutamate receptors abolished all dorsal root‐evoked EPSPs. In comparison to glutamate receptor blockers, SR‐140333 was a weaker antagonist of slow synaptic responses, though it displayed preferential antagonism towards some components of the wind‐up phenomenon. The present results provide evidence obtained with a novel NK1 antagonist that a neuropeptide (presumably substance P), although not directly released by primary afferents onto motoneurons, is a neurotransmitter (acting via NK1 receptors) in the pathway mediating slow synaptic responses of motoneurons, and is presumably involved in signalling nociceptive inputs from the periphery.
Discriminative Cues Indicating Reward Magnitude Continue to Determine Reaction Time of Rats Following Lesions of the Nucleus Accumbens
Authors: Brown Verity J. , Bowman Eric M.
Published: April 2006

The role of the nucleus accumbens in incentive motivation is accepted but poorly understood. In this study, we examined in the rat one aspect of motivated behaviour which might be mediated by the nucleus accumbens, namely the translation of a motivational signal (the expected value of a reward) into motor output (responding for the reward). Rats were trained in a reaction time task in which on each trial they received one, two or three pellets. The number of pellets for each trial was randomly determined in advance and signalled to the rats by cue lights. Rats responded with faster reaction times as the size of the expected reward increased. Following ibotenic acid lesions of the nucleus accumbens, there was no difference in the pattern or the speed of reaction times. Although lesions of the nucleus accumbens did not disconnect the motivational system from the motor system, it is possible that the nucleus accumbens is involved in the learning of the incentive salience of external stimuli. Therefore, after postoperative testing the cue contingencies were reversed. Initially, the cues continued to be interpreted according to their prior significance, but eventually both the lesioned rats and the control group acquired the new relationship and did so in equivalent times. We conclude that the nucleus accumbens is not involved in the acquisition or expression of the processes whereby the expectation of rewards of different value is translated into a motor initiation signal.
Patch‐clamp Analysis of Synaptic Transmission to Cerebellar Purkinje Cells of Prion Protein Knockout Mice
Authors: Herms Jochen W. , Kretzschmar Hans A. , Titz Stefan , Keller Bernhard U.
Published: April 2006

The prion protein (PrP) plays a pivotal role in transmissible spongiform encephalopathies such as Creutzfeldt‐Jakob disease in humans and scrapie and bovine spongiform encephalopathy in animals. Previous experiments have suggested that the normal cellular prion protein (PrPC) is involved in synaptic function in the hippocampus. Here, we utilized the controlled recording conditions of the patch‐clamp technique to investigate the synaptic function of prion protein in cerebellar Purkinje cells. By performing whole‐cell and outside‐out patch‐clamp experiments in thin slices, we investigated synaptic transmission in prion protein knockout mice (PrP‐null) and control animals. In PrP‐null mice, the kinetics of GABA‐ and glutamate receptor‐mediated currents showed no significant deviation from those in control animals. In contrast to previous results in hippocampal neurons, our findings support the view that synaptic transmission is unimpaired in prion protein‐deficient mice.
Metabotropic Glutamate Receptors Inhibiting Excitatory Synapses in the CA1 Area of Rat Hippocampus
Authors: Manzoni Olivier , Bockaert Joël
Published: April 2006

In the CA1 region of hippocampal slices prepared from young adult rats, we studied the ability of several specific agonists of metabotropic glutamate receptors (mGluRs) to depress excitatory synaptic transmission at the CA3–CA1 pyramidal cell synapses. Three groups of mGluRs have been described: group 1 (mGluR1 and 5) receptors are positively coupled to phospholipase C whereas group 2 (mGluR2 and 3) and group 3 (mGluR4, 6, 7 and 8) receptors are negatively coupled to adenylate cyclase. We found that the broad‐spectrum agonist (1S,3R)‐1‐aminocyclopentyl‐1,3‐dicarboxylate and the group 1‐specific agonist (R,S)‐dihydroxyphenylglycine both reversibly inhibited evoked field excitatory postsynaptic potentials, indicating the involvement of group 1 mGluRs. (R,S)‐3,5‐dihydroxyphenylglycine presumably inhibited transmission via a presynaptic mechanism, as whole‐cell voltage‐clamp recordings revealed that inhibition of the synaptic transmission was always accompanied with an increase in paired‐pulse facilitation. Treatment with a specific blocker of mGluR1 receptors, the phenylglycine derivative (S)‐4‐carboxyphenylglycine, was without effect on the (1S,3R)‐1‐amino‐cyclopentyl‐1,3‐dicarboxylate‐induced depression of the field excitatory postsynaptic potentials, strongly suggesting that mGluR5 receptors are responsible for the (1S,3R)‐1‐aminocyclopentyl‐1,3‐dicarboxylate effect. Two selective agonists of group 2 mGluRs, (2S,1′s,2′s)‐2‐(2′‐carboxycyclopropyl)glycine and 4‐carboxy‐3‐hydroxyphenylglycine, were totally ineffective in blocking CA3‐CA1‐evoked synaptic transmission, excluding the involvement of mGluR2/3 subtypes at this developmental stage.
Ischaemia‐induced Long‐term Hyperexcitability in Rat Neocortex
Authors: Luhmann Heiko J. , Mudrick‐Donnon Lori A. , Mittmann Thomas , Heinemann Uwe
Published: April 2006

The long‐term structural and functional consequences of transient forebrain ischaemia were studied with morphological, immunohistochemical and in vitro electrophysiological techniques in the primary somatosensory cortex of Wistar rats. After survival times of 10–17 months postischaemia, neocortical slices obtained from ischaemic animals were characterized by a pronounced neuronal hyperexcitability in comparison with untreated age‐matched controls. Extra‐and intracellular recordings in supragranular layers revealed all‐or‐none long‐latency recurrent responses to orthodromic synaptic stimulation of the afferent pathway. These responses were characterized by durations up to 1.7 s, by multiple components and by repetitive synaptic burst discharges. The reversible blockade of this late activity by dl‐aminophosphonovaleric acid (APV) suggested that this activity was mediated by Kmethyl‐l‐aspartate (NMDA) receptors. The peak conductance of inhibitory postsynaptic potentials was significantly smaller in neurons recorded in neocortical slices obtained from ischaemic animals than those from the controls. However, the average number of parvalbumin (PV)‐labelled neurons per mm3, indicative of a subpopulation of GABAergic interneurons, and the average number and length of dendritic processes arising from PV‐containing cells was not significantly different between ischaemic and control cortex. The prominent dysfunction of the inhibitory system in ischaemic animals occurred without obvious structural alterations in PV‐labelled cells, indicating that this subpopulation of GABAergic interneurons is not principally affected by ischaemia. Our data suggest a long‐term down‐regulation of inhibitory function and a concurrent NMDA receptor‐mediated hyperexcitability in ischaemic neocortex. These alterations may result from structural and/or functional properties of inhibitory non‐PV‐positive neurons or permanent functional modifications on the subcellular molecular level, i.e. alterations in the phosphorylation status of GABA and/or NMDA receptors. The net result of these long‐term changes is an imbalance between the excitatory and inhibitory systems in the ischaemic cortex with the subsequent expression and manifestation of intracortical hyperexcitability.
Initial Tract Formation in the Brain of the Chick Embryo: Selective Expression of the BEN/SC1/DM‐GRASP Cell Adhesion Molecule
Authors: Chédotal Alain , Pourquié Olivier , Sotelo Constantino
Published: April 2006

This study reports the spatio‐temporal pattern of BEN expression (a molecule of the immunoglobulin superfamily) during early stages of the first axonal tract formation, in the fore‐ and midbrain of chick embryos [Hamburger and Hamilton (HH) stages 12–22]. The expression of BEN has been analysed using immunohistochemistry and non‐radioactive in situ hybridization. Furthermore, double labelling experiments (combining anti‐class III β‐tubulin, a pan‐neuronal marker, and anti‐BEN antibodies) have been carried out to determine whether BEN is expressed by all first axonal tracts. The first neurons expressing BEN appear around stage HH13–14, in the caudal diencephalon. They belong to the interstitial nucleus of Cajal, and their axons are the first components of the medial longitudinal fasciculus. By HH14, two other early axonal tracts appear: the tract of the postoptic commissure and the descending root of the mesencephalic nucleus of the trigeminal nerve. Only the latter expresses BEN. At later stages of development numerous new axonal tracts appear in the telencephalic, diencephalic and mesencephalic domains. Only a few of them (the fourth nerve, the lemniscus lateralis, the tectobulbar and habenulopeduncular tracts) express BEN. In all BEN positive systems, the cell bodies, axons and growth cones are uniformly labelled by the antibody. We have found that none of the early axonal tracts grows preferentially at interneuromeric boundaries. Moreover, each tract is formed by several thin fascicles rather than a single one. The expression of BEN is transient and disappears shortly before hatching. These results suggest that BEN may serve to promote axonal outgrowth of precise neuronal systems involved in ‘axonal scaffolding’.
Radiation‐induced Reductions in Macrophage Recruitment Have Only Slight Effects on Myelin Degeneration in Sectioned Peripheral Nerves of Mice
Authors: Perry V. H. , Tsao J. W. , Feam S. , Brown M. C.
Published: April 2006

Macrophage recruitment into the distal nerve stump of the cut or crushed sciatic or saphenous nerves of C57BL/6J mice was reduced by prior whole body irradiation. This procedure was successful in keeping the numbers of cells stained with the mouse macrophage‐specific antibody F4/80 to the levels found in unsectioned nerves. Quantitative image analysis of immunostained sections showed that the rate of loss of myelin basic protein was identical in nerves from irradiated and unirradiated mice up to 5 days but thereafter was slower in macrophage‐deprived nerves. Similar analysis of semithin sections stained with toluidine blue detected more undegenerated myelin in the nerves from irradiated mice 10 days after operation. Quantitative counts made from electron micrographs of the sectioned nerves at 7 days also showed slightly less extensive myelin breakdown in the nerves from irradiated mice. Complete removal of myelin from some Schwann cells can occur without macrophages, but macrophages accelerate the removal of myelin in the later stages of Wallerian degeneration. It is concluded that there are two phases to the breakdown of myelin in peripheral nerves undergoing Wallerian degeneration: an initial stage entirely dependent on the activity of Schwann cells and a later stage dependent on both Schwann cells and the presence of macrophages.