bluuebunny/crossref_metadata_2025 huggingface dataset:
Expression of glutamate transporters in rat optic nerve oligodendrocytes
Authors: Domercq María , Sánchez‐Gómez María Victoria , Areso Pilar , Matute Carlos
Published: March 2003

To investigate the role of glutamate transport in non‐synaptic glia, we characterized the expression of three major glutamate transporters (EAAC1, GLAST and GLT‐1) in rat optic nerve in situ using reverse transcription‐polymerase chain reaction in combination with Western blot and immunochemistry with specific antibodies. GLAST was localized to interfascicular oligodendrocytes, whereas a subpopulation of cells, probably immature oligodendrocyte cells, expressed EAAC1. In contrast, astrocytes, expressed only GLT‐1, consistent with the idea that this is the major glutamate transporter in this cell type. In addition, we observed that glutamine synthetase, a key enzyme in glutamate metabolism, was localized in oligodendrocytes in situ. To examine the properties of these glutamate transporters, we conducted uptake experiments in glial cultures. The kinetics of sodium‐dependent glutamate uptake in cultured oligodendrocytes from the perinatal rat optic nerve were markedly different from those observed in type‐1 astrocytes from the newborn rat cerebral cortex, with higher affinity and lower Vmax. In both cell types, glutamate transport was inhibited by
l‐trans‐pyrrolidine‐2,4‐dicarboxylate (t‐PDC). In contrast, dihydrokainate exhibited significantly more uptake inhibition in oligodendrocytes than in type‐1 astrocytes. These results provide evidence for the expression of functional sodium‐dependent glutamate transporters in optic nerve oligodendrocytes, and suggest that this cell type may play a role in the glutamate–glutamine cycle.
Overexpression of GAP‐43 induces prolonged sprouting and causes death of adult motoneurons
Authors: Harding D. I. , Greensmith L. , Mason M. , Anderson P. N. , Vrbová G.
Published: March 2003

In neurodegenerative diseases, neurons undergo prolonged periods of sprouting. Whether this sprouting compromises these neurons is unknown. Here, we examined the effect of axotomy on adult motoneurons undergoing prolonged sprouting in transgenic mice that overexpress GAP‐43 (growth‐associated protein). Sciatic nerve injury in these adult mice results in motoneuron death, but has no effect in non‐transgenic mice. Thus, continued growth of motor axons renders adult motoneurons susceptible to nerve injury and compromises their long‐term survival. The progressive nature of neurodegenerative diseases may therefore be caused by prolonged sprouting.
Endogenous interleukin‐6 contributes to hypersensitivity to cutaneous stimuli and changes in neuropeptides associated with chronic nerve constriction in mice
Authors: Murphy P. G. , Ramer M. S. , Borthwick L. , Gauldie J. , Richardson P. M. , Bisby M. A.
Published: March 2003

Partial nerve injury is a potential cause of distressing chronic pain for which conventional analgesic treatment with opiates or anti‐inflammatory agents is not very effective. Constriction nerve injury, widely used to study neuropathic pain, was shown here to induce interleukin‐6 (IL‐6) mRNA in a subset of rat primary sensory neurons. When we inflicted chronic nerve constriction on mice with null mutation of the IL‐6 gene, the hypersensitivity to cutaneous heat and pressure that is induced in wild‐type mice was not evident, the loss of substance P in sensory neurons was excessive and the induction of galanin in central sensory projections was reduced. In additional experiments, intrathecal infusion of IL‐6 in rats was shown to stimulate synthesis of galanin in approximately one‐third of lumbar dorsal root ganglion neurons. The results of these experiments indicate that endogenous IL‐6 mediates some of the hypersensitive responses that characterize peripheral neuropathic pain, and influences two neuropeptides that have been implicated in pain transmission.
Regulation of the release of 5‐hydroxytryptamine in the median raphe nucleus of the rat by catecholaminergic afferents
Authors: Adell Albert , Artigas Francesc
Published: March 2003

The present study was conducted in order to examine the influence of catecholaminergic afferents on the release of serotonin in the median raphe nucleus in vivo. To this aim, selective dopamine D1 and D2, and α1‐ and α2‐adrenergic agonists and antagonists were administered locally (1, 10 and 100 μm) through a dialysis probe implanted in the median raphe nucleus of freely moving rats. The D1 and D2 agonists, (±)‐1‐phenyl‐2,3,4,5‐tetrahydro‐(1H)‐3‐benzazepine‐7,8‐diol (SKF‐38393) and quinpirole, respectively, and the D1 and D2 antagonists,
r‐(+)‐7‐chloro‐8‐hydroxy‐3‐methyl‐1‐phenyl‐2,3,4,5‐tetrahydro‐1H‐3‐benzazepine (SCH‐23390) and raclopride, respectively, did not alter the release of serotonin in the median raphe nucleus. The α1‐adrenoceptor agonist phenylephrine did not modify the release of serotonin in this nucleus, although an increased release was observed when the more potent α1‐adrenoceptor agonist cirazoline was used. In contrast, the α1‐adrenoceptor antagonist prazosin reduced the release of 5‐hydroxytryptamine (5‐HT) in a concentration‐dependent manner. The release of 5‐HT was also reduced by the α2‐adrenoceptor agonist clonidine and increased by the α2‐adrenoceptor antagonist 2‐methoxy‐idazoxan (RX821002). These results indicate that the release of serotonin in the median raphe nucleus does not appear to be regulated by dopaminergic afferents through the activation of dopamine D1 or D2 receptors. On the contrary, it is suggested that endogenous noradrenaline exerts a direct tonic stimulatory control on the release of serotonin through α1‐adrenoceptors, and an indirect tonic inhibitory influence through α2‐adrenoceptors located probably in noradrenergic nerve terminals within the median raphe nucleus.
Hu‐Bcl‐2 transgenic mice with supernumerary neurons exhibit timing impairment in a complex motor task
Authors: Rondi‐Reig Laure , Lohof Ann , Dubreuil Yolande Lemaigre , Delhaye‐Bouchaud Nicole , Martinou Jean‐Claude , Caston Jean , Mariani Jean
Published: March 2003

Programmed neuronal cell death is common during development, and is thought to be important in the elimination of errors in axonal projection, cell position and sculpting of neuronal circuits. However, the potential importance of programmed cell death for complex behaviour in the adult animal has never been addressed. We studied motor abilities in a strain of transgenic mice with neuronal overexpression of the human Bcl‐2 protein, which have supernumerary neurons due to reduced developmental cell death. Our results show that these mice have a clear deficiency in fine timing of motor coordination without impairment of basic motor functions. This is the first indication that altered developmental cell death and the consequent neuronal surplus can impair complex behaviour in the adult animal.
Acute application of NGF increases the firing rate of aged rat basal forebrain neurons
Authors: Albeck D. S. , Bäckman C. , Veng L. , Friden P. , Rose G. M. , Granholm A. ‐C. E.
Published: March 2003

Nerve growth factor (NGF) has been widely used in animal models to ameliorate age‐related neurodegeneration, but it cannot cross the blood–brain barrier (BBB). NGF conjugated to an antibody against the transferrin receptor (OX‐26) crosses the BBB and affects the biochemistry and morphology of NGF‐deprived basal forebrain neurons. The rapid actions of NGF, including electrophysiological effects on these neurons, are not well understood. In the present study, two model systems in which basal forebrain neurons either respond dysfunctionally to NGF (aged rats) or do not have access to target‐derived NGF (intraocular transplants of forebrain neurons) were tested. One group of transplanted and one group of aged animals received unconjugated OX‐26 and NGF comixture as a control, while other groups received replacement NGF in the form of OX‐26–NGF conjugate during the 3 months preceding the electrophysiological recording session. Neurons from animals in both the transplanted and aged control groups showed a significant increase in firing rate in response to acute NGF application, while none of the conjugate‐treated groups or young intact rats showed any response. After the recordings, forebrain transplants and aged brains were immunocytochemically stained for the low‐affinity NGF receptor. All conjugate treatment groups showed significantly greater staining intensity compared to controls. These data from both transplants and aged rats in situ indicate that NGF‐deprived basal forebrain neurons respond to acute NGF with an increased firing rate. This novel finding may have importance even for long‐term biological effects of this trophic factor in the basal forebrain.
Neuroendocrine responses to an emotional stressor: evidence for involvement of the medial but not the central amygdala
Authors: Dayas C. V. , Buller K. M. , Day T. A.
Published: March 2003

The amygdala plays a pivotal role in the generation of appropriate responses to emotional stimuli. In the case of emotional stressors, these responses include activation of the hypothalamic–pituitary–adrenal (HPA) axis. This effect is generally held to depend upon the central nucleus of the amygdala, but recent evidence suggests a role for the medial nucleus. In the present study, c‐fos expression, amygdala lesion and retrograde tracing experiments were performed on adult rats in order to re‐evaluate the role of the central as opposed to the medial amygdala in generating neuroendocrine responses to an emotional stressor. Brief restraint (15 min) was used as a representative emotional stressor and was found to elicit c‐fos expression much more strongly in the medial than central nucleus of the amygdala; relatively few Fos‐positive cells were seen in other amygdala nuclei. Subsequent experiments showed that ibotenic acid lesions of the medial amygdala, but not the central amygdala, greatly reduced restraint‐induced activation of cells of the medial paraventricular nucleus, the site of the tuberoinfundibular corticotropin‐releasing factor cells that constitute the apex of the HPA axis. Medial amygdala lesions also reduced the activation of supraoptic and paraventricular nucleus oxytocinergic neurosecretory cells that commonly accompanies stress‐induced HPA axis activation in rodents. To assess whether the role of the medial amygdala in the control of neuroendocrine cell responses to emotional stress might involve a direct projection to such cells, retrograde tracing of amygdala projections to the paraventricular nucleus was performed in combination with Fos immunolabelling. This showed that although some medial amygdala cells activated by exposure to an emotional stressor project directly to the paraventricular nucleus, the number is very small. These findings provide the first direct evidence that it is the medial rather than the central amygdala that is critical to hypothalamic neuroendocrine cell responses during an emotional response, and also provide the first evidence that the amygdala governs oxytocin as well as HPA axis responses to an emotional stressor.
Regionalization of Fos immunostaining in rat accessory olfactory bulb when the vomeronasal organ was exposed to urine
Authors: Inamura Kouhei , Kashiwayanagi Makoto , Kurihara Kenzo
Published: March 2003

The distribution of Fos‐immunoreactive (Fos‐ir) cells in the accessory olfactory bulb (AOB) of rats following vomeronasal organ exposure to urine was studied. Following exposure to male and female Wistar rat urine, Fos‐ir cells were found in the mitral/tufted cell layer, granule cell layer and periglomerular cell layer of the AOB of female Wistar rat, with the highest number in the granule cell layer. Exposure to water or removal of the vomeronasal organ suppressed the expression of Fos‐ir cells. These results suggest that female Wistar rats specifically detect urinary substances derived from male or female Wistar rats via the vomeronasal organ. Exposure of the vomeronasal organ of female Wistar rats to male Wistar urine induced the appearance of many more Fos‐ir cells in all layers of the AOB than exposure to female Wistar urine. As for the mitral/tufted cell layer, the density of Fos‐ir cells in the rostral portion (Gi2α‐positive) of all regions of the AOB was about twice as high as that in the caudal portion when male urine was given. The distribution pattern of Fos‐ir cells in response to female urine was not identical to that in response to male urine. That is, the density of Fos‐ir cells in the caudal portion was slightly larger than that in the rostral portion in the lateral region, while in other regions the density in the rostral portion was higher than that in the caudal portion. It is likely that information from different pheromones is transmitted to the higher brain regions through the different regions of the AOB.
Horizontal cells of the rabbit retina are non‐selectively connected to the cones
Authors: Hack Iris , Peichl Leo
Published: March 2003

Mammalian horizontal cells have generally been assumed to be spectrally non‐selective in their cone contacts until recently, when specific contacts have been found for some species. The rabbit retina is frequently studied as a representative of dichromatic mammalian retinae. These are the reasons for elucidating the connections of the two types of horizontal cells (A‐HCs and B‐HCs) with the green‐sensitive and blue‐sensitive cones of the rabbit retina. Individual A‐HCs and B‐HCs were revealed by Lucifer Yellow injections, the total cone population overlying them was stained using peanut agglutinin, and the blue cones among these were identified by the antiserum JH 455 against blue cone opsin. Both A‐HCs and B‐HCs indiscriminately contact the two cone types available. This holds for the green cone‐dominated dorsal retina and the blue cone‐dominated ventral retina. No evidence was found for a third, potentially blue cone‐selective, horizontal cell type [postulated by Famiglietti, E. V. (1990) Brain Res., 535, 174–179].
Stretch‐activated cation channels of leech neurons: characterization and role in neurite outgrowth
Authors: Calabrese B. , Manzi S. , Pellegrini M. , Pellegrino M.
Published: March 2003

The goal of this study was to characterize the stretch‐activated ion channels (SACs) of adult identified neurons of the leech Hirudo medicinalis and to test the role of SACs in neurite outgrowth of isolated cells. Using cell‐attached patch recording, we established that SACs are densely distributed in the growth cone membrane of cultured neurons. In excised patches, we found that these channels are permeable to Ca2+, as well as to monovalent cations. The channels are blocked by the extracellular application of gadolinium (Gd3+), amiloride and gentamicin. Amiloride and gentamicin, respectively, induce a partial and complete voltage‐dependent block. Time‐lapse video recordings of neurite outgrowth from single cultured neurons were used to study the effects of blocking SACs with gentamicin. Within 20 h of plating in the presence of the aminoglycoside, the total length of neuronal arborization was significantly greater than that measured in its absence. The amount of assembled axon per unitary surface area remained constant over 40 h and did not differ significantly with or without gentamicin. Our findings show that SACs of leech neurons admit Ca2+, are densely distributed in the growth cone membrane and exhibit typical pharmacological features of mechanotransducer ion channels. In addition, our data suggest that these cation channels participate in the early interaction between growing neurites and culture substrate.
Optic flow illusion and single neuron behaviour reconciled by a population model
Authors: Lappe Markus , Duffy Charles J.
Published: March 2003

Radial patterns of optic flow contain a centre of expansion that indicates the observer’s direction of self‐movement. When the radial pattern is viewed with transparently overlapping unidirectional motion, the centre of expansion appears to shift in the direction of the unidirectional motion [Duffy, C.J. & Wurtz, R.H. (1993) Vision Res., 33, 1481–1490]. Neurons in the medial superior temporal (MST) area of monkey cerebral cortex are thought to mediate optic flow analysis, but they do not shift their responses to parallel the illusion created by transparent overlap. The population‐based model of optic flow analysis proposed by Lappe and Rauschecker replicates the illusory shift observed in perceptual studies [Lappe, M. & Rauschecker, J.P. (1995) Vision Res., 35, 1619–1631]. We analysed the behaviour of constituent neurons in the model, to gain insight into neuronal mechanisms underlying the illusion. Single model neurons did not show the illusory shift but rather graded variations of their response specificity. The shift required the aggregate response of the population. We compared the model’s predictions about the behaviour of single neurons with the responses recorded from area MST. The predicted distribution of overlap effects agreed with that observed in area MST. The success of the population‐based model in predicting the illusion and the neuronal behaviour suggests that area MST uses the graded responses of single neurons to create a population response that supports optic flow perception.
Release of endogenous dopamine in cultured mesencephalic neurons: influence of dopaminergic agonists and glucocorticoid antagonists
Authors: Rougé‐Pont Françoise , Abrous Djoher Nora , Le Moal Michel , Piazza Pier Vincenzo
Published: March 2003

Several electrochemical techniques allow the measurement of dopamine release in freely moving animals and brain slices. In this report, we applied one of these techniques, coulometry, coupled to high‐performance liquid chromatography (HPLC), to the study of dopamine release in primary cultures of embryonic mesencephalic dopaminergic neurons. Between day 9 and 33 of culture, concentrations of dopamine, above the detection threshold, were found in the incubation buffer (Krebs ringer buffer, KRB). Concentrations of dopamine in the incubation buffer reflected neuronal release as they were: (i) positively correlated with the number of tyrosine hydroxylase‐positive dopamine neurons in the culture; (ii) tetrodotoxin (TTX) sensitive and Ca2+ dependent; (iii) increased by a depolarizing stimulus, e.g. K+ (20 m
m), or by the indirect dopamine agonists amphetamine and cocaine; (iv) decreased by a hyperpolarizing stimulus, e.g. the dopamine D2‐like receptor agonist quinpirole. Dopamine release in this model was also sensitive to the manipulation of glucocorticoids, potent modulators of dopamine release in vivo. Long‐term treatment of the cell cultures with RU 39305, a selective antagonist of glucocorticoid receptors (GR), but not with spironolactone, a selective antagonist of mineralocorticoid receptors (MR), dose‐dependently decreased K+‐stimulated dopamine release. In conclusion, these results demonstrate an in vitro model that allows the studying of the release of endogenous dopamine in cell cultures and the effects of glucocorticoid hormones on the release dynamics.
Native human neocortex release‐regulating dopamine D2 type autoreceptors are dopamine D2 subtype
Authors: Fedele Ernesto , Fontana Giovanni , Munari Claudio , Cossu Massimo , Raiteri Maurizio
Published: March 2003

Dopamine (DA) autoreceptors expressed at DA nerve terminals regulate DA release. Considerable evidence has indicated that, in rodents, these autoreceptors belong to the D2 type of the DA receptor family, which, in turn, comprises the D2, D3 and D4 subtypes. We investigated here, for the first time, the subclassification of native human DA autoreceptors by studying the release of [3H]DA evoked by electrical stimulation in fresh human neocortical slices. The results have been compared with those obtained in three animal systems: rat neocortical and striatal slices and rat mesencephalic neuronal cultures. In human neocortical slices, the D2/D3 receptor agonist quinpirole (1 n
m–10 μm) inhibited tritium release with a calculated EC50 of 17 n
m and a maximal inhibition of ≈ 75% reached at 1 μm. In the presence of the D2/D3 receptor antagonist (–)‐sulpiride (0.1 and 1 μm), the concentration–response curve of quinpirole was shifted to the right, and the apparent pA2 mean value was 8.5 (8.14–8.77); on the other hand, the inhibitory effects of quinpirole were not affected by the D3 receptor‐selective antagonist [7‐N,N‐dipropylamino‐5,6,7,8‐tetrahydro‐naphtho(2,3b) dihydro,2,3‐furane] (S 14297) and the D4 receptor‐selective antagonist 3‐(4‐[4‐chlorophenyl]piperazin‐1‐yl)‐methyl‐1H‐pyrrolo [2,3‐b]pyridine (L‐745,870) (0.01–1 μm in each case). Superimposable results have been obtained when the release was elicited from rat striatal slices or dopamine mesencephalic neurons in culture, whereas quantitative differences emerged in the case of rat cortical slices. It is concluded that in human brain, as well as in rat brain, the release of DA in the terminal region of midbrain dopaminergic neurons is regulated through autoreceptors of the D2 subtype.
Reduced hippocampal LTP in mice lacking a presynaptic protein: complexin II
Authors: Takahashi Sei‐ichi , Ujihara Hisamitsu , Huang Guang‐Zhe , Yagyu Ken‐ichi , Sanbo Makoto , Kaba Hideto , Yagi Takeshi
Published: March 2003

The SNAP receptor (SNARE) complex is a core complex specialized for synaptic vesicle exocytosis, and the binding of SNAPs to the complex is an essential step for neurotransmitter release. Complexin I and II have been identified as SNARE‐complex‐associated proteins. Importantly, complexins compete with α‐SNAP for binding to the complex, suggesting that complexins may modulate neurotransmitter release process. To examine this possibility and to understand the physiological function of complexins, we generated complexin II knockout mice. The complexin‐II‐deficient mice (–/–) were viable and fertile, and appeared normal. Electrophysiological recordings in the mutant hippocampus showed that ordinary synaptic transmission and paired‐pulse facilitation, a form of short‐term synaptic plasticity, were normal. However, long‐term potentiation (LTP) in both CA1 and CA3 regions was impaired, suggesting that complexin II may not be essential for synaptic vesicle exocytosis, but it does have a role in the establishment of hippocampal LTP.
Calpain activation and inhibition in organotypic rat hippocampal slice cultures deprived of oxygen and glucose
Authors: Brana C. , Benham C. D. , Sundstrom L. E.
Published: March 2003

It has been suggested that, after ischaemia, activation of proteases such as calpains could be involved in cytoskeletal degradation leading to neuronal cell death. In vivo, calpain inhibitors at high doses have been shown to reduce ischaemic damage and traumatic brain injury, however, the relationship between calpain activation and cell death remains unclear. We have investigated the role of calpain activation in a model of ischaemia based on organotypic hippocampal slice cultures using the appearance of spectrin breakdown products (BDPs) as a measure of calpain I activation. Calpain I activity was detected on Western blot immediately after a 1‐h exposure to ischaemia. Up to 4 h post ischaemia, BDPs were found mainly in the CA1 region and appeared before uptake of the vital dye propidium iodide (PI). 24 h after the insult, BDPs were detected extensively in CA1 and CA3 pyramidal cells, all of which was PI‐positive. However, there were many more PI‐positive cells that did not have BDPs, indicating that the appearance of BDPs does not necessarily accompany ischaemic cell death. Inhibition of BDP formation by the broad‐spectrum protease inhibitor leupeptin was not accompanied by any neuroprotective effects. The more specific and more cell‐permeant calpain inhibitor MDL 28170 had a clear neuroprotective effect when added after the ischaemic insult. In contrast, when MDL 28170 was present throughout the entire pre‐ and post‐incubation phases, PI labelling actually increased, indicating a toxic effect. These results suggest that calpain activation is not always associated with cell death and that, while inhibition of calpains can be neuroprotective under some conditions, it may not always lead to beneficial outcomes in ischaemia.
Inhibition of glial scarring in the injured rat brain by a recombinant human monoclonal antibody to transforming growth factor‐β2
Authors: Logan Ann , Green Jonathan , Hunter Allison , Jackson Ronald , Berry Martin
Published: March 2003

The transforming growth factor‐βs (TGF‐βs) are potent fibrogenic factors implicated in numerous central nervous system (CNS) pathologies in which fibrosis and neural dysfunction are causally associated. In this study, we aim to limit the fibrogenic process in a model of CNS scarring using a recombinant human monoclonal antibody, derived from phage display libraries and specific to the active form of the TGF‐β2 isoform. The implicit inference of the work was that, as such antibodies are potential pharmacological agents for the treatment of human CNS fibrotic diseases, validation of efficacy in a mammalian animal model is a first step towards this end. Treatment of cerebral wounds with the anti‐TGF‐β2 antibody led to a marked attenuation of all aspects of CNS scarring, including matrix deposition, formation of an accessory glial‐limiting membrane, inflammation and angiogenesis. For example, in the wound, levels of: (i) the connective tissue components fibronectin, laminin and chondroitin sulphate proteoglycan; and (ii) wound‐responsive cells including astrocytes and macrophages/microglia, were markedly reduced. Our findings suggest that such synthetic anti‐fibrotic TGF‐β antibodies are potentially applicable to a number of human CNS fibrotic diseases to arrest the deposition of excessive extracellular matrix components, and maintain and/or restore functional integrity.
Postnatal development of GABAB receptor‐mediated modulation of voltage‐activated Ca2+ currents in mouse brain‐stem neurons
Authors: Zhang W. , Elsen F. , Barnbrock A. , Richter D. W.
Published: March 2003

GABAB receptors modulate respiratory rhythm generation in adult mammals. However, little is currently known of their functional significance during postnatal development. In the present investigation, the effects of GABAB receptor activation on voltage‐activated Ca2+ currents were examined in rhythmically active neurons of the pre‐Bötzinger complex (PBC). Both low‐ (LVA) and high‐voltage‐activated (HVA) Ca2+ currents were present from the first postnatal day (P1). The density of LVA Ca2+ currents increased during the first week, whilst the density of HVA Ca2+ currents increased after the first week. In the second postnatal week, the HVA Ca2+ currents were composed of L‐ (47 ± 10%) and N‐type (21 ± 8%) currents plus a ‘residual’ current, whilst there were no N‐type currents detectable in the first few days. The GABAB receptor agonist baclofen (30 μm) increased LVA Ca2+ currents (30 ± 11%) at P1–P3, but it decreased the currents (35 ± 11%) at P7–P15 without changing its time course. At all ages, baclofen (30 μm) decreased the HVA Ca2+ currents by ≈ 54%. Threshold of baclofen effects on both LVA and HVA Ca2+ currents was 5 μm at P1–P3 and lower than 1 μm at P7–P15. The effect of baclofen was abolished in the presence of the GABAB receptor antagonist CGP 55845A (50 n
m). We conclude that both LVA and HVA Ca2+ currents increased postnatally. The GABAB receptor‐mediated modulation of these currents undergo marked developmental changes during the first two postnatal weeks, which may contribute essentially to modulation of respiratory rhythm generation.
Modulation of steroidal levels by adrenalectomy/castration and inhibition of neurosteroid synthesis enzymes affect σ1 receptor‐mediated behaviour in mice
Authors: Phan Vân‐Ly , Su Tsung‐Ping , Privat Alain , Maurice Tangui
Published: March 2003

The interaction between neurosteroids and sigma1 (σ1) receptors may be of therapeutic interest during physiological or pathological ageing, particularly concerning their neuromodulatory role on cognitive functions. Neurosteroids modulate memory processes through a mechanism involving interactions with GABAA, N‐methyl‐
d‐aspartate and/or σ1 receptors. To measure the contribution of endogenous neurosteroid levels to the antiamnesic effects of σ1 agonists, we investigated the effects of inhibitors of key enzymes involved in neurosteroid synthesis, in adrenalectomized/castrated (AdX/CX) mice to avoid the effect of circulating steroids. Trilostane, a 3β‐hydroxysteroid‐deshydrogenase inhibitor, blocks the pregnenolone to progesterone conversion and leads to a decrease of progesterone. Finasteride, a 5α‐reductase inhibitor, blocks the progesterone to 5α‐pregnane‐3,20‐dione conversion and leads to an accumulation of progesterone. The in vivo binding of (+)‐[3H]SKF‐10 047 to σ1 sites was measured in the mouse hippocampus and cortex. The attenuating effect of the selective σ1 agonist PRE‐084 (0.1–3 mg/kg) against dizocilpine (0.15 mg/kg)‐induced learning impairment was examined using spontaneous alternation behaviour, step‐down passive avoidance and place learning in the elevated plus‐maze. The in vivo (+)‐[3H]SKF‐10 047 binding appeared significantly increased in AdX/CX mice and after trilostane treatment (10 mg/kg twice a day, 7 days), compared with sham‐operated animals. The finasteride treatment (25 mg/kg, 7 days) significantly decreased binding levels. The learning deficits induced by dizocilpine were not affected by the treatments. The antiamnesic effect of PRE‐084 was facilitated in AdX/CX mice and even more after trilostane treatment, as several parameters for animals treated with both PRE‐084 and dizocilpine returned to control values. The PRE‐084 effect was blocked after finasteride. These results confirmed that endogenous neurosteroidal levels modulate σ1 receptor‐mediated behaviour directly, and revealed that, among neurosteroids, progesterone may be the main modulator of σ1 receptors.
Li+ and muscarine cooperatively enhance the cationic tail current in rat cortical pyramidal cells
Authors: Okada Takashi , Kang Youngnam , Ohmori Harunori
Published: March 2003

Li+ is known to facilitate the onset of status epilepticus induced by cholinergic stimulation, although the underlying mechanisms are not clear. Under whole‐cell current clamp conditions with a CsCl‐based internal solution, cortical pyramidal cells display a single plateau‐spike followed by a slow depolarizing afterpotential (DAP) in response to injection of a short current pulse. However, the same current pulse generated a burst of plateau‐spikes after application of Li+ (2 m
m) and muscarine (10 μm). As similar bursts of plateau‐spikes were generated through an enhancement of the slow DAP when [K + ]o was raised (
Kang et al. 1998
), we have investigated the effects of Li+ and muscarine on the Ca2+‐dependent cationic current underlying the slow DAP, measured as the slow tail current evoked after the offset of depolarizing voltage pulses. Muscarine enhanced the amplitudes of both early and late components of the slow tail current. This effect of muscarine was markedly potentiated by Li+, while Li+ by itself affected the slow tail current only slightly. Intracellular application of heparin (0.5–1 mg/mL) suppressed the effect of muscarine in the presence of Li+. These results suggest that inositol‐trisphosphate‐induced Ca2+ release is involved in the cooperative enhancement of the slow tail current, and this cooperation may be one of the mechanisms underlying facilitation of the onset of epilepsy induced by these agents.
Neuroprotection of cultured foetal rat hippocampal cells against glucose deprivation: are GABAergic neurons less vulnerable or more sensitive to TCP protection?
Authors: Chaudieu Isabelle , Privat Alain
Published: March 2003

In the rat brain, hippocampal neurons are particularly sensitive to secondary excitotoxic injury induced by ischaemia or hypoglycaemia. To determine some distinctive features of vulnerability among neuronal phenotypes in the hippocampus following a metabolic insult, we used an in vitro model of mild glucose deprivation. Primary cultures from the rat hippocampus (21 days in vitro) were deprived of glucose for 4 h and then were returned to the standard medium for 24 or 48 h. Survival of the GABAergic neuronal population was evaluated both by measuring [3H]GABA uptake and by counting GAD65‐immunostained cells. This was compared with the survival of the total neuronal population evaluated by counting the neurofilament‐200‐immunostained cells. Glucose deprivation for 4 h followed by a recovery period of 48 h induced a decrease of 59% and 40% in the number of GAD65‐ and neurofilament‐200‐immunostained cells, respectively. Thus, GABAergic neurons were slightly more vulnerable to glucose deprivation than the other neurons in the hippocampal cell cultures. When the excitotoxic component of cellular death was blocked in the presence of TCP, an NMDA‐antagonist, the survival of GABAergic neurons was almost complete after 48 h of recovery. In contrast, measurements of the release of lactate dehydrogenase in the medium indicated that TCP largely protected hippocampal cells after 24 h but was ineffective after 48 h. This observation was confirmed by immunostaining data which showed that after 48 h TCP did not significantly increase the survival of neurofilament‐200‐immunostained cells. These results indicate that after glucose deprivation and a recovery period of 48 h, GABAergic neurons in hippocampal cell cultures are not more resistant than other neurons but they are more sensitive to TCP protection.
Expression in mammalian cells and electrophysiological characterization of two mutant Kv1.1 channels causing episodic ataxia type 1 (EA‐1)
Authors: Bretschneider Frank , Wrisch Anja , Lehmann‐Horn Frank , Grissmer Stephan
Published: March 2003

Episodic ataxia type 1 (EA‐1) is a rare neurological disorder and was the first ionic channel disease to be associated with defects in a potassium channel. Until now 10 different point mutations in the KCNA1‐gene have been reported to cause this disorder. We have investigated the functional consequences of two mutations leading to amino acid substitutions in the first and sixth transmembrane segments of a Kv1.1 channel subunit, by means of the patch‐clamp technique; we injected cRNA coding for, respectively, F184C and V408A mutant Kv1.1 channels into mammalian cells and compared the resulting currents with those in the wild‐type. The expression levels of F184C and V408A mutant channels relative to that of the wild‐type was 38 and 68%, respectively. Since the single‐channel conductance of the F184C mutant was similar to that of the wild‐type (12 pS) without an apparent change in the maximum open probability, we conclude that the lower expression level in the F184C mutant channels is due to a reduced number of functional channels on the cell surface. F184C activated slower, and at more depolarized potentials, and deactivated faster compared with the wild‐type. V408A channels deactivated and inactivated faster compared with the wild‐type. Studies with different extracellular cations and tetraethylammonium gave no indication that the pore structure was changed in the mutant channels. Acetazolamide, that is helpful in some patients suffering from EA‐1, was without effect on Kv1.1 wild‐type or mutant channels. This study confirms and extends earlier studies on the functional consequences of Kv1.1 mutations associated with EA‐1, in an attempt to understand the pathophysiology of the disease.
Can GABAA conductances explain the fast oscillation frequency of absence seizures in rodents?
Authors: Destexhe Alain
Published: March 2003

Rodent models of absence epilepsy generate spike‐and‐wave oscillations at relatively fast frequency (5–10 Hz) compared with humans (≈ 3 Hz). Possible mechanisms for these oscillations were investigated by computational models that included the complex intrinsic firing properties of thalamic and cortical neurons, as well as the multiple types of synaptic receptors mediating their interactions. The model indicates that oscillations with spike‐and‐wave field potentials can be generated by thalamocortical circuits. The frequency of these oscillations critically depended on GABAergic conductances in thalamic relay cells, ranging from 2–4 Hz for strong GABAB conductances to 5–10 Hz when GABAA conductances were dominant. This model therefore suggests that thalamocortical circuits can generate two types of spike‐and‐wave oscillations, whose frequency is determined by the receptor type mediating inhibition in thalamic relay cells. Experiments are proposed to test this mechanism.
Facilitation of GABA release by arachidonic acid in rat hippocampal synaptosomes
Authors: Cunha R. A. , Ribeiro J. A.
Published: March 2003

Arachidonic acid (AA) is proposed to be a facilitatory retrograde messenger in hippocampal glutamatergic synapses. In this study, we found that AA (10 μM) increased the basal outflow (19 ± 4%) and the K+‐evoked release of [3H]GABA (38 ± 3%) from rat hippocampal synaptosomes. This effect is likely to be a direct action of AA, as it was not mimicked by arachidic acid (10 μ M) and was not modified by inhibition of either lipooxygenase with nordihydroguaiaretic acid (50 μM) or cyclooxygenase with indomethacin (100 μM). Activation of protein kinase C may be involved, as chelerythrine (6 μM), a protein kinase C inhibitor, attenuated the AA (10 μM)‐facilitation of K+‐evoked [3H]GABA release by 58 ± 5%. Phospholipase A2 (2 U/mL), an enzyme that releases AA, and melittin (1 μM), a phospholipase A2 activator, mimicked the AA‐facilitation of evoked [3H]GABA release (70 ± 6% and 76 ± 7% facilitation, respectively). These results show that exogenously added and endogenously produced AA increased basal outflow and K+‐evoked release of [3H]GABA from rat hippocampal synaptosomes. Thus, AA can no longer be considered solely a facilitatory neuromodulator in the hippocampus, as this AA‐facilitation of the release of the main inhibitory neurotransmitter may predominate under certain circumstances.
The intrinsic specification of γ‐aminobutyric acid type A receptor α6 subunit gene expression in cerebellar granule cells
Authors: Bahn Sabine , Wisden William , Dunnett Stephen B. , Svendsen Clive
Published: March 2003

The patterns of γ‐aminobutyric acid type A (GABAA) receptor subunit gene expression in the brain are complex. For example, mouse hippocampal dentate granule cells express many subunit genes, whereas adult cerebellar granule cells, which may share differentiation mechanisms, have a smaller compliment and uniquely express the α6 subunit gene. To see how the α6 expression component arises, i.e. if intrinsically or environmentally specified, we used a mouse line (Δα6lacZ) with a β‐galactosidase reporter inserted into the α6 gene. Precursor cells from postnatal day 1 Δα6lacZ cerebellum were transplanted to the adult hippocampus and cerebellum of wild‐type mice; 4 weeks after transplantation, Δα6lacZ cells expressed α6‐lacZ in the hippocampus, amygdala and cerebellum. Thus, different adult environments support both the development and maintenance of α6 gene expression from cerebellar granule cell precursors. Establishing α6 gene expression is not likely to require specific patterns of neurotransmitter innervation or other factors present only in the developing brain; instead, α6 expression can be timed and maintained autonomously.
Selective expression of the NOS II isoform during mouse vestibulocochlear receptorgenesis
Authors: Arnhold S. , Klinz F. J. , Bloch W. , Hess A. , Andressen C. , Addicks K.
Published: March 2003

Based on in vitro studies, nitric oxide (NO) is reported to be involved in initial neuronal differentiation. In order to compare this finding with the situation in vivo, we have looked for the expression of the three NO synthase isoforms in the developing mouse vestibulocochlear system. From these isoforms only the inducible NOS II is expressed during inner ear development. Examination of a series of embryonic and early postnatal animals, up to postnatal day 6, reveals a maturation‐dependent, monophasic expression of this isoform. Initial expression is observed by day 10 of gestation in nerve cells of the vestibolocochlear ganglion and on their fibres. By day 14 of gestation, these afferent fibres penetrate the epithelium of the prospective receptor fields making contact with early, differentiating immunoreactive cochlear hair cells and receptor cells of the macula and crista ampullaris. This receptor‐cell‐derived immunoreactivity vanished in differentiated sensory hair cells by postnatal day 6, when both the constitutive isoforms and subsequent activated members of the down stream second messenger cascade (guanylate cyclase/cGMP) of the adult mouse were not then detectable. The strict phasic expression of NOS‐II, independent of the second messenger system mentioned above, implies that there is a unique role for the inducible NOS isoform in nerve cell differentiation, independent of the NO/guanylate cyclase/cGMP pathway.
Calcium‐dependent inactivation of the monosynaptic NMDA EPSCs in rat hippocampal neurons in culture
Authors: Medina Igor , Leinekugel Xavier , Ben‐Ari Yehezkel
Published: March 2003

The effects of increased dendritic calcium concentration ([Ca2+]i) induced by single action potentials on monosynaptic glutamatergic excitatory postsynaptic currents (EPSCs) were studied in cultured rat hippocampal neurons. To investigate the respective roles of pre‐ and postsynaptic elements in the depolarization‐induced NMDAR inactivation, we have performed simultaneous paired whole‐cell recordings from monosynaptically connected pre‐ and postsynaptic hippocampal neurons. We report that the single firing of the postsynaptic neuron did not result in inactivation of the NMDAR‐EPSC, whereas a burst of depolarizing steps transiently depressed the NMDAR‐EPSCs in both pyramidal cells and interneurons. This effect was mediated by postsynaptic voltage‐gated Ca2+ influx, as it was prevented by: (i) buffering postsynaptic [Ca2+]i with 30 m
m BAPTA; (ii) removing extracellular Ca2+; or (iii) applying Cd2+o (100 μm), a voltage‐gated calcium channel blocker. It does not involve presynaptic mechanisms as it selectively affected NMDA but not α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionate (AMPA) receptor‐mediated EPSCs. These results suggest that inactivation of NMDAR‐channels by voltage‐gated Ca influx is a general property of hippocampal neurons, which may play an important role in reducing postsynaptic NMDAR Ca2+ influx that leads to plasticity or excitotoxicity during sustained neuronal activity.
Behavioural conditions affecting saccadic eye movements elicited electrically from the frontal lobes of primates
Authors: Tehovnik Edward J. , Slocum Warren M. , Schiller Peter H.
Published: March 2003

We assessed the effects of varying the time at which electrical stimulation was delivered to the dorsomedial frontal cortex (DMFC) and the frontal eye fields (FEF) relative to the onset of a visual target. Monkeys were required to fixate the visual target to obtain a drop of apple juice as reward. We found that the probability of eliciting saccades increased with increases in the delay of electrical stimulation relative to target onset. Also, the current threshold to evoke saccades decreased as electrical stimulation was delivered later following target onset. There were major differences in the magnitude of this effect with stimulation of the DMFC versus the FEF. The current threshold to evoke saccades from the DMFC was 16 times greater when electrical stimulation was delivered 200 ms after target onset as compared to when it was delayed 200 ms after target offset. In contrast, the current threshold to evoke saccades from the FEFs was only three times greater when stimulation was delivered under similar conditions. These results suggest that the FEF are more closely connected with the saccade generator for the execution of saccadic eye movements than is the DMFC, even though both regions have direct projections to brainstem oculomotor centres.
Long‐term effects of methylprednisolone following transection of adult rat spinal cord
Authors: Oudega M. , Vargas C. G. , Weber A. B. , Kleitman N. , Bunge M. B.
Published: March 2003

Clinically, high‐dose treatment with the glucocorticosteroid, methylprednisolone (MP), within 8 h after spinal cord injury, has been shown to improve neurological recovery. The current standard of care is to administer MP as a bolus of 30 mg/kg followed by a 23‐h infusion of 5.4 mg/kg/h to spinal cord injured patients. To better understand the role of MP in neuroprotection, we have studied how MP administration affects macrophage accumulation, tissue loss, and axonal dieback at 1, 2, 4 and 8 weeks after a complete transection of the eighth thoracic spinal cord in the adult rat. A 30 mg/kg dose of MP was administered intravenously at 5 min, and 2 and 4 h after injury. The number of ED1 (antibody against microglia/macrophages) ‐positive cells was quantified in a 500‐μm‐wide strip of tissue directly adjacent and parallel to the transection. At all time points, MP treatment led to a significant decrease in the number of ED1‐positive cells in both rostral and caudal stumps. Over the 2‐month post‐transection period, the average MP‐induced reduction in the number of ED1‐positive cells was 82% in the rostral cord stump and 66% in the caudal stump. Using a computerized image analysis system, it was observed that MP treatment resulted in a significant reduction in tissue loss in both cord stumps at 2, 4 and 8 week post‐injury. Over the 2‐month post‐lesion period, the average MP‐induced reduction in tissue loss in the caudal cord stump was higher than that in the rostral stump; 48 versus 37%, respectively. Immunostaining for neurofilaments and growth‐associated protein‐43 (GAP‐43) revealed the presence of numerous axons near and in the lesion site. Anterograde neuronal tracing with biotinylated dextran amine showed that, in MP‐treated animals, dieback of vestibulospinal fibres, but not of corticospinal fibres, was significantly diminished at all time points studied. In addition, with MP administration, 1 and 2 weeks after injury, an increase in the number of vestibulospinal fibres was found at 1 and 2 mm from the transection, suggesting transient regenerative sprouting of these fibres. The results demonstrate that treatment with MP shortly after spinal cord transection in the adult rat led to a long‐term reduction of ED1‐positive cells and spinal tissue loss, reduced dieback of vestibulospinal fibres, and a transient sprouting of vestibulospinal fibres near the lesion at 1 and 2 weeks post‐lesion. The possible relationships between the inflammatory changes, spinal tissue sparing, and axonal survival and sprouting are complex and need to be further explored.