bluuebunny/crossref_metadata_2025 huggingface dataset:
Metabolic coupling between glia and neurons is necessary for maintaining respiratory activity in transverse medullary slices of neonatal mouse
Authors: Hülsmann Swen , Oku Yoshitaka , Zhang Weiqi , Richter Diethelm W.
Published: March 2003

The respiratory rhythm is generated and regulated by a neuronal network within the lower brainstem. While the neuronal mechanisms of rhythm generation have been extensively investigated, the contribution of glial cells remains to be determined. Here we report the effect of specific blockade of the glial Krebs cycle and glutamine synthetase on the neuronal activity of the respiratory network. Application of 5 mm fluoroacetate, which selectively blocks the glial Krebs cycle, suppressed rhythmic respiratory burst activity. Substitution of either the Krebs cycle substrate isocitrate (3 mm) or glutamine (3 mm) restored rhythmic network activity. Blockade of glutamine synthetase by methionine sulfoximine (0.5 mm) suppressed rhythmic burst activity as well. Resubstitution of glutamine (3 mm) was able to restore rhythmic activity in the presence of methionine sulfoximine. This data demonstrates that the glutamate–glutamine cycle in astrocytes and their supply of glutamine to neuronal glutamatergic terminals is essential for the rhythm generation in the respiratory centre.
Location of calcium transporters at presynaptic terminals
Authors: Juhaszova Magdalena , Church Paul , Blaustein Mordecai P. , Stanley Elis F.
Published: March 2003

The plasma membrane ATP‐driven Ca2+ pump (PMCA) and the Na+/Ca2+ exchanger (NCX) are the major means of Ca2+ extrusion at presynaptic nerve terminals, but little is know about the location of these transporters relative to the major sites of Ca2+ influx, the transmitter release sites. We used immunocytochemistry to identify these transport proteins in a calyx‐type presynaptic nerve terminal from the ciliary ganglion of the chick. The PMCA clusters were localized to the transmitter release sites, as identified by staining for the secretory vesicle‐specific protein synaptotagmin I. This colocalization was not due to the presence of the pump on the secretory vesicle itself because membrane fractionation of chick brain synaptosomes demonstrated comigration of the pump with surface membrane and not vesicle markers. In contrast, the NCX did not colocalize with synaptotagmin but tended to be located at nonsynaptic regions of the terminal. The PMCA location, near the transmitter release sites, suggests that it plays a role in priming the release site by maintaining a low free Ca2+ level, facilitating the dissociation of the ion from its binding sites. The PMCA may also replenish external Ca2+ in the synaptic cleft following periods of synaptic activity. In contrast, the NCX location suggests a role in the rapid emptying of cytoplasmic Ca2+ uptake organelles which serve as the main line of defence against high free Ca2+.
Differential effects of NT‐3 on reinnervation of the fast extensor digitorum longus (EDL) and the slow soleus muscle of rat
Authors: Simon M. , Terenghi G. , Green C. J. , Coulton G. R.
Published: March 2003

Previous studies of gastrocnemius muscle reinnervation showed specific normalization of the proportion and diameter of fast type 2b muscle fibres following NT‐3 delivery to the proximal stump of the cut sciatic nerve. Here, we investigate if normalization was related to greater improvement of muscle reinnervation of fast (extensor digitorum longus; EDL) than slow (soleus) motor units. NT‐3‐impregnated (NT‐3 group) or plain fibronectin (FN group) mats were inserted into a sciatic nerve gap. Neuromuscular junctions (NMJs) labelled with TRITC‐α‐bungarotoxin were colabelled with calcitonin gene‐related peptide (CGRP) or 4E2 antisera and imaged using confocal microscopy. CGRP and 4E2 were used as markers for newly reinnervated and structurally mature NMJs, respectively. At 40 days postsurgery, denervated NMJs in EDL and soleus muscles of both groups presented a 50% decrease of surface area due to decreased width. At day 80 in EDL, more NMJs were reinnervated by CGRP‐immunoreactive terminals in the NT‐3 (7.1%) than in the FN group (4.2%); there was no difference between groups for soleus. At 120 days, 4E2‐immunoreactive NMJs were more numerous in EDL of the NT‐3 (40.0%) than in the FN group (7.3%), unlike in soleus (NT‐3, 1.6%; FN, 1.8%), and presented a partial size recovery. These results indicate that NT‐3 preferentially improves reinnervation of fast muscles over slow muscle, although the mechanism of this improvement is still unclear.
Enlarged cholinergic forebrain neurons and improved spatial learning in p75 knockout mice
Authors: Greferath Ursula , Bennie Antoinette , Kourakis Angela , Bartlett Perry F. , Murphy Mark , Barrett Graham L.
Published: March 2003

The p75 low affinity neurotrophin receptor (p75) can induce apoptosis in various neuronal and glial cell types. Because p75 is expressed in the cholinergic neurons of the basal forebrain, p75 knockout mice may be expected to show an increased number of neurons in this region. Previous studies, however, have produced conflicting results, suggesting that genetic background and choice of control mice are critical. To try to clarify the conflicting results from previous reports, we undertook a further study of the basal forebrain in p75 knockout mice, paying particular attention to the use of genetically valid controls. The genetic backgrounds of p75 knockout and control mice used in this study were identical at 95% of loci. There was a small decrease in the number of cholinergic basal forebrain neurons in p75 knockout mice at four months of age compared with controls. This difference was no longer apparent at 15 months due to a reduction in numbers in control mice between the ages of 4 and 15 months. Cholinergic cell size in the basal forebrain was markedly increased in p75 knockout mice compared with controls. Spatial learning performance was consistently better in p75 knockout mice than in controls, and did not show any deterioration with age. The results indicate that p75 exerts a negative influence on the size of cholinergic forebrain neurons, but little effect on neuronal numbers. The markedly better spatial learning suggests that the function, as well as the size, of cholinergic neurons is negatively modulated by p75.
Formation of hippocampal synapses on patterned substrates of a laminin‐derived synthetic peptide
Authors: Matsuzawa Mieko , Tabata Toshihide , Knoll Wolfgang , Kano Masanobu
Published: March 2003

We created a new culture system which provides simple, stereotyped neuronal circuitries suitable for investigating synaptic events between mammalian central neurons. We used surface chemistry and laser‐lithography to produce geometrical patterns of neuron‐compatible substrate spaced by less neuron‐compatible surfaces. The patterned substrates were composed of a laminin‐derived synthetic peptide, PA22‐2, and the spacing surfaces of either decyldimethylsilane (DDMS) or trimethylsilane (TMS). Dissociated rat hippocampal neurons survived on the patterned substrates for several days without the aid of glia and extended their neurites along the substrates. The TMS spacing surfaces appeared more favourable for the excitability development and axonal differentiation of the hippocampal neurons, but less favourable for the development of the resting conductance than the DDMS spacing surfaces. Furthermore, neurons grown on the patterned substrates frequently made synaptophysin‐positive contacts with one another. Spontaneous post‐synaptic currents recorded from such neurons suggest that these contacts were indeed functional synapses. When hippocampal neurons were plated at a very low density, they often formed circuitries consisting of only two neurons on the patterned substrate. Such a simple circuitry allowed us to analyse synaptic transmission in a single neuronal pair without the influence of the third neurons. With the clarity of analysis and the readiness of manipulation, our culture system would offer a powerful tool for studying development and functions of mammalian central synapses.
Survival effects of BDNF and NT‐3 on axotomized rubrospinal neurons depend on the temporal pattern of neurotrophin administration
Authors: Novikova Liudmila N. , Novikov Lev N. , Kellerth Jan‐Olof
Published: March 2003

This study shows that both BDNF and NT‐3 can prevent cell death in axotomized adult rat rubrospinal neurons (RSNs), but that the efficacy of neuroprotection depends on the temporal pattern of treatment. At 8 weeks after cervical spinal cord injury, 51% of the RSNs had died. Subarachnoidal BDNF infusion into the cisterna magna for 4 weeks resulted in neuronal hypertrophy and 71% survival. Continuous infusion for 8 weeks into the lumbar subarachnoidal space with either BDNF or NT‐3 gave similar survival rates, while a combination of BDNF and NT‐3 resulted in 96% survival, although the cells were atrophic. When administration of either BDNF or NT‐3 was delayed and performed during postoperative weeks 5–8, the number of surviving neurons was increased compared to early treatment. Delayed treatment with a combination of BDNF and NT‐3 resulted in complete survival and a reduction in neuronal atrophy. A decreased expression of TrkB receptors and microtubule‐associated protein‐2 in the RSNs after axotomy was counteracted by BDNF and NT‐3. Microglial activity remained increased even when complete cell survival was achieved. Thus, the combination of neurotrophins as well as the temporal pattern of treatment need to be adequately defined to optimize survival of injured spinal tract neurons.
Cyclooxygenase‐1 is a marker for a subpopulation of putative nociceptive neurons in rat dorsal root ganglia
Authors: Chopra B. , Giblett S. , Little J. G. , Donaldson L. F. , Tate S. , Evans R. J. , Grubb B. D.
Published: March 2003

Immunocytochemical and morphometric techniques were used to quantify the distribution of cyclooxygenase (cox)‐containing neurons in rat L5 dorsal root ganglia (DRG). Cox‐1 immunolabelling was almost exclusively restricted to small diameter DRG neurons (< 1000 μm2), and was extensively colocalized with calcitonin gene‐related peptide (CGRP) and isolectin B4 (IB4). Cox‐1 was present in 65% and 70% of CGRP‐ and IB4‐labelled neurons, respectively. Cox‐1 labelling was also found in neurons expressing the sensory neuron‐specific (SNS) Na+ channel. Cox‐2 labelling was absent in DRG from normal rats. In the Freund's adjuvant model of monoarthritis, the proportion of cox‐1‐positive DRG neurons was unchanged and no neurons were found to be labelled for cox‐2. In primary tissue culture, cox‐1 immunolabelling persisted in vitro for up to 9 days and was present in morphologically identical neurons. The selective expression of cox‐1 in peripheral ganglia was confirmed by the small number of nodose ganglion neurons and superior cervical ganglion (SCG) neurons labelled for cox‐1. These data suggest that cox‐1 is a marker for a subpopulation of putative nociceptive neurons in vitro and in vivo, and suggests that the prostaglandins synthesized by these neurons may be important for nociceptor function. These data may have important implications for the mode and mechanism of action of non‐steroidal anti‐inflammatory drugs (NSAIDs).
Developmental and adult expression of rat calcium‐sensing receptor transcripts in neurons and oligodendrocytes
Authors: Ferry Sandrine , Traiffort Elisabeth , Stinnakre Jacques , Ruat Martial
Published: March 2003

The calcium‐sensing receptor (CaSR) is a member of a growing family of heptahelical receptors with an unusually large extracellular domain. To further delineate its functions in neurons and glia, we have investigated the expression pattern of CaSR transcripts in the postnatal and adult rat brain, spinal cord and dorsal root ganglia by in situ hybridization. CaSR‐expressing cells were spatially and temporally regulated in myelinated structures with a caudo‐rostral pattern that paralleled that of myelin basic protein, a marker of myelination, with a downregulation observed in the adult. Double‐labelling studies demonstrated that CaSR mRNA colocalizes with myelin basic protein‐expressing cells within fibre tracts, suggesting that CaSR is expressed by mature oligodendrocytes. In cultured rat oligodendrocytes, Ca2+ induced stimulation of phosphatidylinositol hydrolysis with an EC50 of 1.4 mm and increased intracellular calcium. NPS R‐568 (1 μm), a calcimimetic, significantly stimulates the inositol phosphate response, whereas a less potent stereoisomer, NPS S‐568 (1 μm), was without effect. These data suggest that a functional CaSR is expressed in mature oligodendrocytes with a potential role in myelination. CaSR expression was also developmentally regulated in neurons of the orbital cortex and in the CA2 region of the hippocampus, and present in olfactory nuclei, hypothalamic areas and in the area postrema through postnatal days to adulthood. This expression is consistent with a role of CaSR in olfactory or gustatory signal integration, and with the regulation of fluid and mineral homeostasis. CaSR expression in a subpopulation of small cells in dorsal root ganglia suggests additional roles for extracellular Ca2+ in sensory nerves.
Acidic motif responsible for plasma membrane association of the voltage‐dependent calcium channel β1b subunit
Authors: Bogdanov Y. , Brice N. L. , Canti C. , Page K. M. , Li M. , Volsen S. G. , Dolphin A. C.
Published: March 2003

Voltage‐dependent calcium channels consist of a pore‐forming transmembrane α1‐subunit, which is known to associate with a number of accessory subunits, including α2‐δ‐ and β‐subunits. The β‐subunits, of which four have been identified (β1–4), are intracellular proteins that have marked effects on calcium channel trafficking and function. In a previous study, we observed that the β1b‐subunit showed selective plasma membrane association when expressed alone in COS7 cells, whereas β3 and β4 did not. In this study, we have examined the basis for this, and have identified, by making chimeric β‐subunits, that the C‐terminal region, which shows most diversity between β‐subunits, of β1b is responsible for its plasma membrane association. Furthermore we have identified, by deletion mutations, an 11‐amino acid motif present in the C terminus of β1b but not in β3 (amino acids 547–556 of β1b, WEEEEDYEEE), which when deleted, reduces membrane association of β1b. Future research aims to identify what is binding to this sequence in β1b to promote membrane association of this calcium channel subunit. It is possible that such membrane association is important for the selective localization or clustering of particular calcium channels with which β1b is associated.
Mirtazapine enhances frontocortical dopaminergic and corticolimbic adrenergic, but not serotonergic, transmission by blockade of α2‐adrenergic and serotonin2C receptors: a comparison with citalopram
Authors: Millan M. J. , Gobert A. , Rivet J. ‐M. , Adhumeau‐Auclair A. , Cussac D. , Newman‐Tancredi A. , Dekeyne A. , Nicolas J. ‐P. , Lejeune F.
Published: March 2003

Mirtazapine displayed marked affinity for cloned, human α2A‐adrenergic (AR) receptors at which it blocked noradrenaline (NA)‐induced stimulation of guanosine‐5′‐O‐(3‐[35S]thio)‐triphosphate ([35S]‐GTPγS) binding. Similarly, mirtazapine showed high affinity for cloned, human serotonin (5‐HT)2C receptors at which it abolished 5‐HT‐induced phosphoinositide generation. Alpha2‐AR antagonist properties were revealed in vivo by blockade of UK‐14,304‐induced antinociception, while antagonist actions at 5‐HT2C receptors were demonstrated by blockade of Ro 60 0175‐induced penile erections and discriminative stimulus properties. Mirtazapine showed negligible affinity for 5‐HT reuptake sites, in contrast to the selective 5‐HT reuptake inhibitor, citalopram. In freely moving rats, in the dorsal hippocampus, frontal cortex (FCX), nucleus accumbens and striatum, citalopram increased dialysate levels of 5‐HT, but not dopamine (DA) and NA. On the contrary, mirtazapine markedly elevated dialysate levels of NA and, in FCX, DA, whereas 5‐HT was not affected. Citalopram inhibited the firing rate of serotonergic neurons in dorsal raphe nucleus, but not of dopaminergic neurons in the ventral tegmental area, nor adrenergic neurons in the locus coeruleus. Mirtazapine, in contrast, enhanced the firing rate of dopaminergic and adrenergic, but not serotonergic, neurons. Following 2 weeks administration, the facilitatory influence of mirtazapine upon dialysate levels of DA and NA versus 5‐HT in FCX was maintained, and the influence of citalopram upon FCX levels of 5‐HT versus DA and NA was also unchanged. Moreover, citalopram still inhibited, and mirtazapine still failed to influence, dorsal raphe serotonergic neurons. In conclusion, in contrast to citalopram, mirtazapine reinforces frontocortical dopaminergic and corticolimbic adrenergic, but not serotonergic, transmission. These actions reflect antagonist properties at α2A‐AR and 5‐HT2C receptors.
Inhibition of delayed rectifier K+ conductance in cultured rat cerebellar granule neurons by activation of calcium‐permeable AMPA receptors
Authors: Jones Glyn , Boyd David F. , Yeung Shuk Yin , Mathie Alistair
Published: March 2003

Activation of AMPA (α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole propionic acid) receptors in cerebellar granule cells during perforated‐patch whole‐cell recordings activated an inward current at negative voltages which was followed, after a delay, by the inhibition of an outward potassium current at voltages positive to −20 mV. The activated inward current was inwardly rectifying suggesting that the AMPA receptors were Ca2+‐permeable. This was confirmed by direct measurements of intracellular calcium where Ca2+ rises were seen following AMPA receptor activation in Na+‐free external solution. Ca2+ rises were equally large in the presence of 100 μm Cd2+ to block voltage‐gated Ca2+ channels. Specific voltage‐protocols, allowing selective activation of the delayed rectifier potassium current (KV) and the transient A current (KA), showed that kainate inhibited KV, but not to any great extent KA. The inhibition of KV was blocked by the AMPA receptor antagonist CNQX (6‐cyano‐7‐nitroquinoxaline‐2,3‐dione) and was no longer observed when the KV current was abolished with high concentrations of Ba2+. The responses to kainate were not altered by pre‐treating the cells with pertussis toxin, suggesting that the AMPA receptor stimulation of the G‐protein Gi cannot account for the effects observed. Replacing extracellular Na+ with choline did not alter the inhibition of KV by kainate, however, removing extracellular Ca2+ reduced the kainate response. The inhibition of KV by kainate was unaffected by the presence of 100 μm Cd2+. The guanylyl cyclase inhibitor, ODQ (1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one), did not alter kainate inhibition of KV. It is concluded that ion influx (particularly Ca2+ ions) through AMPA receptor channels following receptor activation leads to an inhibition of KV currents in cerebellar granule neurons.
Cell death is prevented in thalamic fields but not in injured neocortical areas after permanent focal ischaemia in mice overexpressing the anti‐apoptotic protein Bcl‐2
Authors: De Bilbao Fabienne , Guarin Ernesto , Nef Patrick , Vallet Philippe , Giannakopoulos Panteleimon , Dubois‐Dauphin Michel
Published: March 2003

Previous studies have suggested that various apoptotic‐related proteins could be involved in the death process induced by cerebral ischaemia. In order to further clarify their role and examine how the anti‐apoptotic protein Bcl‐2 could influence this process, the time‐course of mRNA expression of various cell death genes was studied from 1 to 14 days following permanent occlusion of the middle cerebral artery in wild‐type (WT) and Bcl‐2 transgenic mice, within and outside the area of infarction. No differences of the infarct sizes were observed between the two groups of mice, showing that the extent of neuronal injury could not have been lowered by the Bcl‐2 transgene. Seven days after the ischaemic insult, the mRNA expression of the cell death gene effector cpp32 was dramatically upregulated in the penumbra of WT and Bcl‐2 transgenic mice. Interestingly, the cpp32 transcript was markedly induced from 3 days in the ipsilateral thalamus of the two groups of mice. However, apoptotic bodies were observed in the thalamic field of WT but not transgenic mice. This suggests that cpp32 mRNA may be induced in an attempt to kill the injured cells and, in contrast to the penumbra, cell death in the thalamus may be prevented in Bcl‐2 transgenic mice. Based on these results, the pathophysiological mechanisms that underly neuronal damage following ischaemia need consideration in order to evaluate the extent of neuroprotection that may be afforded by the Bcl‐2 anti‐apoptotic protein. Although the present study does not confirm previous data showing a protective role of Bcl‐2 in neocortical infarcted areas, it suggests that anti‐apoptotic therapies may constitute a possible treatment for areas of the brain remote from those directly affected by ischaemia.
Induction of integrin‐associated protein (IAP) mRNA expression during memory consolidation in rat hippocampus
Authors: Lee Eminy H. Y. , Hsieh Yi P. , Yang Chia L. , Tsai Kuen J. , Liu Cheng H.
Published: March 2003

We have previously demonstrated that hippocampal integrin‐associated protein (IAP) gene expression is associated with memory formation in a one‐way inhibitory avoidance learning in rats. In the present study, we further investigated the role and mechanism of IAP involved in memory consolidation in rats. Because of the minute amount of IAP present in the brain, we have adopted the quantitative reverse transcription‐polymerase chain reaction (RT‐PCR) method. Our results revealed that hippocampal IAP mRNA expression is approximately fourfold higher in rats showing good memory retention (GM, retention score of 600 s) at 3 h, but not at other time points, after training when compared with the poor memory rats (retention score < 80 s). On the other hand, integrin αv mRNA level was markedly increased (∼ twofold), while integrin β3 mRNA level was decreased (∼ 50%) at 1 h post‐training. Further, separate sets of RT‐PCR analysis revealed that IAP5 and IAP6 mRNA expressions, but not that of IAP7, were markedly increased in GM rats 3 h post‐training. Moreover, regional distribution studies revealed that different isoforms of the IAP gene are similarly distributed in different brain areas, while IAP7 has been the predominant form present in astrocyte cells. These results together suggest that IAP mRNA expression is indeed induced upon training, rather than that the GM rats have constitutively higher levels of IAP. The unparallel change of IAP and integrin mRNA expressions as far as time‐course is concerned suggests that they are possibly involved in different forms and stages of memory processing. Further, IAP5 and IAP6 are more closely associated with memory consolidation, while IAP7 may constitute the major isotype for signal transduction in astrocyte cells.
Alterations in Purkinje cell spines of calbindin D‐28 k and parvalbumin knock‐out mice
Authors: Vecellio Marco , Schwaller Beat , Meyer Michael , Hunziker Willi , Celio Marco R.
Published: March 2003

The second messenger Ca2+ is known to act in a broad spectrum of fundamental cell processes, including modifications of cell shape and motility, through the intermediary of intracellular calcium‐binding proteins. The possible impact of the lack of the intracellular soluble Ca2+‐binding proteins parvalbumin (PV) and calbindin D‐28 k (CB) was tested on spine morphology and topology in Purkinje cell dendrites of genetically modified mice. Three different genotypes were studied, i.e. PV or CB single knock‐out (PV–/–, CB–/–) and PV and CB double knock‐out mice (PV–/–CB–/–). Purkinje cells were microinjected with Lucifer Yellow and terminal dendrites scanned at high resolution with a confocal laser microscope followed by three‐dimensional (3‐D) reconstruction. The absence of PV had no significant effect on spine morphology, whereas the absence of CB resulted in a slight increase of various spine parameters, most notably spine length. In double knock‐out mice, the absence of both PV and CB entailed a doubling of spine length, an increase in spine volume and spine surface, a higher spine density along the dendrites, as well as a more clustered spine distribution. In all three genotypes, a reduction in the number of stubby spines was observed compared with wild‐type animals. These results suggest a morphological compensation for the lack of the soluble calcium buffers in the cytoplasm of Purkinje cell dendritic spines. The increase in various spine parameters, particularly volume, may counteract the lack of the calcium buffers, such as to adjust Ca2+‐transients at the transitional zone between spines and dendrites.
Red nucleus lesions impair overground locomotion in rats: a kinetic analysis
Authors: Muir Gillian D. , Whishaw Ian Q.
Published: March 2003

The red nucleus is a prominent brainstem nucleus in mammals which is thought to be involved in production of skilled limb movements. The presence of the red nucleus and associated rubrospinal tract in animals that do not produce skilled limb movements, however, suggests that these structures might also be involved in control of more general limb actions, such as those occurring during locomotion. The present study investigates this question by measuring the three‐dimensional ground reaction forces produced by locomoting rats with unilateral excitotoxic lesions of the red nucleus. Twenty‐four to 48 h after the lesion, rats moved with an asymmetric gait during which abnormal braking and propulsive forces were produced during the dual contact time of the forelimb contralateral to the lesion and the ipsilateral hindlimb. Rats did not recover normal symmetrical locomotion within the 55‐day duration of the study. The persistent asymmetry produced by red nucleus ablation provides the first unequivocal demonstration that the red nucleus plays a role in ongoing overground locomotion in the rat. Species differences in phylogeny and connectivity of the red nucleus are discussed, as well as the possibility that there is a general compensatory response to unilateral CNS injury in the rat.
Alterations in preproenkephalin and adenosine‐2a receptor mRNA, but not preprotachykinin mRNA correlate with occurrence of dyskinesia in normal monkeys chronically treated with l‐DOPA
Authors: Zeng B. ‐Y. , Pearce R. K. B. , MacKenzie G. M. , Jenner P.
Published: March 2003

Chronic treatment with l‐DOPA induces dyskinesia in patients with Parkinson's disease (PD) and 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP)‐treated monkeys, but is not thought to do so in normal humans or primates. However, we have shown that chronic oral high dose l‐DOPA administration, with the peripheral decarboxylase inhibitor, carbidopa and with or without the peripherally acting catechol‐O‐methyl transferase (COMT) inhibitor, entacapone, to normal macaque monkeys for 13 weeks induced dyskinesia in a proportion of animals. In the present study, in situ hybridization histochemistry was used to investigate the effect of chronic l‐DOPA administration on the activity of the direct and indirect striatal output pathways by measuring striatal preprotachykinin (PPT), preproenkephalin‐A (PPE‐A) and adenosine‐2a (A2a) receptor gene expression in these monkeys. Overall there was no significant difference in striatal PPT, PPE‐A and A2a receptor mRNA levels between normal animals and all l‐DOPA (plus carbidopa and/or entacapone)‐treated animals irrespective of whether or not dyskinesia occurred. However, when the level of PPE‐A and A2a receptor mRNA was analysed in eight monkeys displaying marked dyskinesias as a result of l‐DOPA (plus carbidopa with or without entacapone) treatment, there was a significant increase in PPE‐A and A2a receptor mRNA message levels in the striatum compared with animals receiving identical treatment, but displaying few or no involuntary movements, and compared with normal controls. There was no difference in striatal PPT mRNA levels in monkeys exhibiting severe dyskinesia compared with those showing little or no dyskinesia after l‐DOPA treatment or to normal controls. These results suggest that prolonged l‐DOPA treatment alone has no consistent effect on either the direct or indirect pathways, as judged by striatal PPT, PPE‐A or A2a receptor mRNA levels in normal monkeys. However, in monkeys exhibiting marked dyskinesia resulting from chronic l‐DOPA treatment, abnormal activity is detected in the indirect striato‐pallidal output pathway, as judged by striatal PPE‐A and A2a receptor mRNA levels, indicating an imbalance between the direct and indirect striatal pathway which may explain the emergence of dyskinesia in these animals.
Molecular and functional properties of synaptically activated NMDA receptors in neonatal motoneurons in rat spinal cord slices
Authors: Abdrachmanova Galya , Teisinger Jan , Vlachová Viktorie , Vyklický Ladislav
Published: March 2003

The functional properties of N‐methyl‐ d‐aspartate (NMDA) receptor‐mediated excitatory postsynaptic currents (EPSC) were studied in fluorescence‐labelled motoneurons in thin spinal cord slices. The deactivation of NMDA receptor EPSCs in motoneurons voltage‐clamped at +40 mV was independent of intensity or location of stimulation and of postnatal age [τfast = 28.5 ± 4.6 ms (63.6 ± 8.8%) and τslow = 165.6 ± 49.6 ms]. In the presence of 1 mm Mg2+ the amplitude of NMDA receptor EPSCs was voltage‐dependent. Boltzmann analysis of the relationship between peak NMDA receptor EPSC amplitude and membrane potential suggested an apparent Kd of Mg2+ (at 0 mV) of 0.87 mm. Nonstationary variance analysis of NMDA receptor EPSCs gave an estimated single‐channel conductance of 59 ± 14 pS. Direct measurement of the NMDA receptor channel openings in outside‐out patches isolated from motoneurons indicated the presence of single‐channel conductance levels of 21.8 ± 2.8 pS, 37.1 ± 3.2 pS, 49.6 ± 5.1 pS and 69.6 ± 4.2 pS. Single‐cell RT‐PCR analysis of mRNA revealed that NR1, NR2A–D and NR3A transcripts were expressed in motoneurons. These results suggest that specific assembly of NMDA receptor subunits in motoneurons determines the functional and pharmacological properties of the receptors in these cells.
Transcription factor dynamics and neuroendocrine signalling in the mouse pineal gland: a comparative analysis of melatonin‐deficient C57BL mice and melatonin‐proficient C3H mice
Authors: Von Gall Charlotte , Lewy Alfred , Schomerus Christof , Vivien‐Roels Berthe , Pevét Paul , Korf Horst‐Werner , Stehle Jörg H.
Published: March 2003

In rodents, the nocturnal rise and fall of arylalkylamine N‐acetyltransferase (AANAT) activity controls the rhythmic synthesis of melatonin, the hormone of the pineal gland. This rhythm involves the transcriptional regulation of the AANAT by two norepinephrine (NE)‐inducible transcription factors, e.g. the activator pCREB (phosphorylated Ca2+/cAMP‐response element binding protein) and the inhibitor ICER (inducible cAMP early repressor). Most inbred mouse strains do not produce melatonin under standard laboratory light/dark conditions. As melatonin‐deficient mice are often the founders for transgenic animals used for chronobiological experimentations, molecular components of neuroendocrine signalling in the pineal gland as an integral part of clock entrainment mechanisms have to be deciphered. We therefore compared calcium signalling, transcriptional events and melatonin synthesis in the melatonin‐deficient C57BL mouse and the melatonin‐proficient C3H mouse. Pineal glands and primary pinealocytes were cultured and stimulated with NE or were collected at various times of the light/dark (LD) cycle. Changes in intracellular calcium concentrations, the phosphorylation of CREB, and ICER protein levels follow similar dynamics in the pineal glands of both mouse strains. pCREB levels are high during the early night and ICER protein shows elevated levels during the late night. In the C57BL pineal gland, a low but significant increase in melatonin synthesis could be observed upon NE stimulation, and, notably, also when animals were exposed to long nights. We conclude that the commonly used C57BL mouse is not completely melatonin‐deficient and that this melatonin‐deficiency does not affect molecular details involved in regulating transcriptional events of melatonin synthesis.
Tissue plasminogen activator controls multiple forms of synaptic plasticity and memory
Authors: Calabresi Paolo , Napolitano Maddalena , Centonze Diego , Marfia Girolama A. , Gubellini Paolo , Teule Martine Ammassari , Berretta Nicola , Bernardi Giorgio , Frati Luigi , Tolu Massimo , Gulino Alberto
Published: March 2003

Induction of long‐term depression (LTD) in rat striatal slices revealed that this form of synaptic plasticity is coupled to an increased expression of tissue‐plasminogen activator (t‐PA) mRNA, as detected by the mRNA differential display technique. To further investigate the involvement of this gene in synaptic remodelling following striatal LTD, we recorded electrical activity from mice lacking the gene encoding t‐PA (t‐PA‐KO) and from wild‐type (WT) mice. Tetanic stimulation induced LTD in the large majority of striatal neurons recorded from WT mice. Conversely, LTD was absent in a significant proportion of striatal neurons obtained from mice lacking t‐PA. Electrophysiological recordings obtained from hippocampal slices in the CA1 area showed that mainly the late phase of long‐term potentiation (LTP) was reduced in t‐PA‐KO mice. Learning and memory‐related behavioural abnormalities were also found in these transgenic mice. Disruption of the t‐PA gene, in fact, altered both the context conditioning test, a hippocampus‐related behavioural task, and the two‐way active avoidance, a striatum‐dependent task. In an open field object exploration task, t‐PA‐KO mice expressed deficits in habituation and reactivity to spatial change that are consistent with an altered hippocampal function. Nevertheless, decreased rearing and poor initial object exploration were also observed, further suggesting an altered striatal function. These data indicate that t‐PA plays a critical role in the formation of various forms of synaptic plasticity and memory.
Effect of nitric oxide and NO synthase inhibition on nonquantal acetylcholine release in the rat diaphragm
Authors: Mukhtarov M. R. , Urazaev A. KH. , Nikolsky E. E. , Vyskočil F.
Published: March 2003

After anticholinesterase treatment, the postsynaptic muscle membrane is depolarized by about 5 mV due to nonquantal release of acetylcholine (ACh) from the motor nerve terminal. This can be demonstrated by the hyperpolarization produced by the addition of curare (H‐effect). The magnitude of the H‐effect was decreased significantly to 3 mV when the nitric oxide (NO) donors, sodium nitroprusside (SNP) and S‐nitroso‐N‐acetylpenicillamine (SNAP) were applied to the muscle, or when NO production was elevated by addingl‐arginine, but notd‐arginine, as a substrate. The H‐effect was increased to 8–9 mV by inhibition of NO synthase byl‐nitroarginine methylester (l‐NAME), or by guanylyl cyclase inhibition by methylene blue and 1H‐[1,2,4]oxidiazolo[4,3‐a]quinoxalin‐1‐one (ODQ). ODQ increased the H‐effect to 7.3 ± 0.2 mV and diminished the SNP‐induced decrease of the H‐effect when applied together with SNP. The effects of NO donors andl‐arginine were eliminated by adding reduced haemoglobin, an extracellular NO scavenger. The present results, together with earlier evidence for the presence of NO synthase in muscle fibres, indicate that nonquantal release of ACh is modulated by NO production in the postsynaptic cell.
Glycine receptors containing the α4 subunit in the embryonic sympathetic nervous system, spinal cord and male genital ridge
Authors: Harvey Robert J. , Schmieden Volker , Von Holst Alexander , Laube Bodo , Rohrer Hermann , Betz Heinrich
Published: March 2003

Inhibitory glycine receptors (GlyRs) are known to mediate postsynaptic inhibition in spinal cord, brain stem and some higher brain regions. Several developmentally and regionally regulated GlyR isoforms exist, which result from a differential expression of the GlyR α (α1‐α4) and β subunit genes. Currently, very little is known about GlyRs containing the α4 subunit, whose existence was predicted from a partial genomic sequence. Here, we describe the isolation of complementary DNA (cDNA) sequences for the mouse and chick GlyR α4 subunits. We show that a mouse GlyR α4 subunit full‐length cDNA directs the formation of functional homo‐oligomeric strychnine‐sensitive GlyRs in Xenopus laevis oocytes and mammalian cells, and that these resemble GlyRs composed of the α1 subunit in pharmacological profile and single‐channel properties. In situ hybridization reveals high levels of GlyR α4 subunit transcripts in the embryonic (E13) chick spinal cord, lumbosacral sympathetic ganglia and dorsal root ganglia. The avian GlyR α4 subunit gene also shows male‐specific expression in the developing genital ridge. The pharmacological profile of α4 subunit‐containing receptors and deduced location of the avian GlyR α4 subunit are consistent with it being a component of the embryonic excitatory GlyRs previously identified in sympathetic neurons. Our data also suggest a novel role for GlyRs in the maturation of reproductive organs.
The retrograde inhibition of IPSCs in rat cerebellar Purkinje cells is highly sensitive to intracellular Ca2+
Authors: Glitsch Maike , Parra Paula , Llano Isabel
Published: March 2003

The Ca2+‐dependent retrograde inhibition of inhibitory postsynaptic currents (depolarization‐induced‐suppression of inhibition; DSI) was investigated using fura‐2 Ca2+ measurements and whole‐cell patch‐clamp recordings in rat cerebellar Purkinje cells. DSI was studied in cells loaded with different concentrations of the Ca2+ chelators BAPTA and EGTA. A concentration of 40 mm BAPTA was required to significantly interfere with DSI, whereas 10 mm BAPTA was almost ineffective. 40 mm EGTA reduced DSI, but was less effective than 40 mm BAPTA. Ratiometric Ca2+ measurements indicated that the extent of DSI depended critically on the changes in intracellular calcium ([Ca2+]i). The relationship between DSI and peak Δ[Ca2+]i could be approximated by a hyperbolic function, with apparent half‐saturation concentrations of 200 and 40 nm for dendritic and somatic [Ca2+]i, respectively. It is suggested that DSI is due to somatodendritic exocytosis of a retrograde messenger, and that this exocytosis is highly sensitive to [Ca2+]i.
Does the migraine aura reflect cortical organization?
Authors: Dahlem M. A. , Engelmann R. , Löwel S. , Müller S. C.
Published: March 2003

Individuals suffering from classical migraine report an astonishing diversity of migraine auras. A frequently reported symptom is a visual hallucination known as fortification illusion (FI). Here we demonstrate that the typical zig‐zag pattern of the FI can be reproduced using experimental data of orientation maps of the primary visual cortex (V1) assuming that a continuous excitation front propagates across V1. We put forward a model in which the cortical neurons within this excitation wave are activated sufficiently to contribute to conscious perception. It is shown that the discontinuous repetitive nature of the zig‐zag pattern of the FI can reflect the specific layout of visual cortical orientation maps. Additionally, dynamic features of the FI are predicted based on our model.
The dopaminergic hyper‐responsiveness of the shell of the nucleus accumbens is hormone‐dependent
Authors: Barrot Michel , Marinelli Michela , Abrous Djoher Nora , Rougé‐Pont Françoise , Le Moal Michel , Piazza Pier Vincenzo
Published: March 2003

The dopaminergic projection to the shell of the nucleus accumbens is the most reactive to stress, reward and drugs of abuse and this subregion of the nucleus accumbens is also considered a target of therapeutic effects of atypical antipsychotic drugs (APD). In this report we show, by means of in vivo microdialysis and Fos immunohistochemistry, that the hyper‐responsiveness which characterizes the dopaminergic transmission to the shell is dependent on glucocorticoid hormones. In Sprague‐Dawley rats, after suppression of endogenous glucocorticoids by adrenalectomy, extracellular dopamine levels selectively decreased in the shell, whilst they remained unchanged in the core. This effect was observed in basal conditions, after a mild stress (vehicle injection), as well as after subcutaneous administration of morphine (2 mg/kg, s.c.) or intraperitoneal injection of cocaine (15 mg/kg, i.p.). The decrease in dopamine observed in the shell had a postsynaptic impact, as shown by less induction of Fos‐like proteins selectively in the shell in response to cocaine. However, the induction of Fos‐like proteins by the full D1 agonist SKF82958 (1.5 mg/kg, i.p.) remained unchanged after adrenalectomy, suggesting that the changes in Fos expression after cocaine injection were likely to depend on changes in extracellular dopamine levels rather than on changes in postsynaptic sensitivity to dopamine. The effects of adrenalectomy were glucocorticoid‐specific given that they were prevented by corticosterone treatment. This anatomical specificity in the control of neuronal activity by a hormonal input highlights the role of steroid hormones in shaping the functional activity of the brain.
Depolarizing stimulation upregulates GA‐binding protein in neurons: a transcription factor involved in the bigenomic expression of cytochrome oxidase subunits
Authors: Zhang Chenyang , Wong‐Riley Margaret T. T.
Published: March 2003

Neurons are unique in having dendrites that extend far away from their cell bodies. Mitochondria located in the dendrites can be separated from the nucleus for long distances. The mechanism of bigenomic coordination is of particular importance to cytochrome oxidase (CO), which has subunits that are encoded in both the nuclear and mitochondrial DNA. GA‐binding protein (GABP) is a transcription factor that is required for the promoter activity of mitochondrial transcription factor A as well as several nuclear‐encoded CO subunits. Thus, GABP may play a key role in coordinating the transcription of both mitochondrial and nuclear‐encoded subunits of CO. The goal of the present study was to determine if GABP was expressed in neurons and whether and how it responded to increased neuronal activity. Using primary neuronal cultures, the β‐subunit of GABP was localized immunocytochemically to both the cytoplasm and the nucleus, whereas the α‐subunit was expressed mainly in the nucleus. In KCl‐treated cultures, immunoreactivity for both α‐ and β‐subunits was significantly increased in the nucleus compared with untreated sister cultures. The induction of GABP preceded that of CO gene expression from the two genomes, which, in turn, preceded that of CO activity. Thus, our data suggest that neuronal activity regulates subunit concentrations of GABP in the nucleus, and GABP may be a critical sensor of changes in neuronal activity. Our data are also consistent with the postulated role of GABP as a coordinator of both mitochondrial and nuclear transcription for subunits of CO in neurons.
Outgrowth‐promoting molecules in the adult hippocampus after perforant path lesion
Authors: Savaskan Nicolai E. , Skutella Thomas , Bräuer Anja U. , Plaschke Martina , Ninnemann Olaf , Nitsch Robert
Published: March 2003

Lesion‐induced neuronal plasticity in the adult central nervous system of higher vertebrates appears to be controlled by region‐ and layer‐specific molecules. In this study we demonstrate that membrane‐bound hippocampal outgrowth‐promoting molecules, as present during the development of the entorhino‐hippocampal system and absent or masked in the adult hippocampus, appear 10 days after transection of the perforant pathway. We used an outgrowth preference assay to analyse the outgrowth preference of axons from postnatal entorhinal explants on alternating membrane lanes obtained from hippocampus deafferented from its entorhinal input taken 4, 10, 20, 30 and 80 days post‐lesion and from adult control hippocampus. Neurites from the entorhinal cortex preferred to extend axons on hippocampal membranes disconnected from their entorhinal input for 10 days in comparison with membranes obtained from unlesioned adult animals. Membranes obtained from hippocampi disconnected from their entorhinal input for 10 days were equally as attractive for growing entorhinal cortex (EC) axons as membranes from early postnatal hippocampi. Further analysis of membrane properties in an outgrowth length assay showed that entorhinal axons extended significantly longer on stripes of lesioned hippocampal membranes in comparison with unlesioned hippocampal membranes. This effect was most prominent 10 days after lesion, a time point at which axonal sprouting and reactive synaptogenesis are at their peak. Phospholipase treatment of membranes obtained from unlesioned hippocampi of adult animals strongly promoted the outgrowth length of entorhinal axons on these membranes but did not affect their outgrowth preference for deafferented hippocampal membranes. Our results indicate that membrane‐bound outgrowth‐promoting molecules are reactivated in the adult hippocampus following transection of the perforant pathway, and that neonatal entorhinal axons are able to respond to these molecules. These findings support the hypothesis of a temporal accessibility of membrane‐bound factors governing the layer‐specific sprouting of remaining axons following perforant path lesion in vivo.
Bicuculline injections into the rostral and caudal motor thalamus of the monkey induce different types of dystonia
Authors: Guehl D. , Burbaud P. , Boraud T. , Bioulac B.
Published: March 2003

The pathophysiology of dystonia remains unclear in comparison with other movement disorders. Recent data suggest that there may exist in dystonia an increased thalamic drive to the mesial premotor cortex. To test this hypothesis, we induced overactivity of the motor thalamus by injecting a GABA‐A (γ‐aminobutyric acid) antagonist (bicuculline) into the rostral (pallidal) and caudal (cerebellar) ventrolateral nuclei of the thalamus in both hemispheres of one monkey. Dystonic postures were observed in the contralateral limbs and axis. Electromyographic recordings revealed bursts of muscular activation with co‐contractions during spontaneous dystonic movements and alterations in muscular patterns during sequential visually guided arm movements. The type of dystonia depended on the site of injections. Rostral thalamic injections induced more severe dystonic postures, whereas myoclonic jerks predominated following caudal injections. We conclude that these two distinct clinical patterns, which are frequently associated in humans, are probably due to a dysfunctioning of segregated thalamic projections to the supplementary motor area (from the rostral part) and to the primary motor cortex (from the caudal part).