bluuebunny/crossref_metadata_2025 huggingface dataset:
Role of retinoids in the CNS: differential expression of retinoid binding proteins and receptors and evidence for presence of retinoic acid
Authors: Zetterström Rolf H. , Lindqvist Eva , De Urquiza Alexander Mata , Tomac Andreas , Eriksson Ulf , Perlmann Thomas , Olson Lars
Published: March 2003

Retinoic acid (RA), a retinoid metabolite, acts as a gene regulator via ligand‐activated transcription factors, known as retinoic acid receptors (RARs) and retinoid X receptors (RXRs), both existing in three different subtypes, α, β and γ. In the intracellular regulation of retinoids, four binding proteins have been implicated: cellular retinol binding protein (CRBP) types I and II and cellular retinoic acid binding protein (CRABP) types I and II. We have used in situ hybridization to localize mRNA species encoding CRBP‐ and CRABP I and II as well as all the different nuclear receptors in the developing and adult rat and mouse central nervous system (CNS), an assay to investigate the possible presence of RA, and immunohistochemistry to also analyse CRBP I‐ and CRABP Iimmunoreactivity (IR). RXRβ is found in most areas while RARα and ‐β and RXRα and ‐γ show much more restricted patterns of expression. RARα is found in cortex and hippocampus and RARβ and RXRγ are both highly expressed in the dopamine‐innervated areas caudate/putamen, nucleus accumbens and olfactory tubercle. RARγ could not be detected in any part of the CNS. Using an in vitro reporter assay, we found high levels of RA in the developing striatum. The caudate/putamen of the developing brain showed strong CRBP I‐IR in a compartmentalized manner, while at the same time containing many evenly distributed CRABP I‐IR neurons. The CRBP I‐ and CRABP I‐IR patterns were closely paralleled by the presence of the corresponding transcripts. The specific expression pattern of retinoid‐binding proteins and nuclear retinoid receptors as well as the presence of RA in striatum suggests that retinoids are important in many brain structures and emphasizes a role for retinoids in gene regulatory events in postnatal and adult striatum.
Substance P release in the dorsal horn assessed by receptor internalization: NMDA receptors counteract a tonic inhibition by GABAB receptors
Authors: Marvizón Juan Carlos G. , Grady Eileen F. , Stefani Enrico , Bunnett Nigel W. , Mayer Emeran A.
Published: March 2003

Inhibitory amino acids have antinociceptive actions in the spinal cord that may involve inhibition of neurotransmitter release from primary afferents. Rat spinal cord slices with dorsal roots were used to study the effect of GABA and glycine on substance P release, assessed by the internalization of neurokinin 1 receptors. After electrical stimulation of the dorsal root at 100 Hz, about half of neurokinin 1 receptor‐immunoreactive neurons in laminae I–IIo showed internalization. This internalization was inhibited by GABA (100 μm) and the GABAB agonist R‐baclofen (10 μm), but not by the GABAA agonist muscimol (20 μm) or glycine (100 μm). The GABAB antagonist 2‐hydroxysaclofen (100 μm) reversed the inhibitory effect of GABA, but not the GABAA antagonist bicuculline (100 μm). These findings demonstrate that GABAB receptors, but not GABAA or glycine receptors, inhibit substance P release induced by dorsal root stimulation. In contrast, R‐baclofen did not inhibit the internalization produced by NMDA (100 μm), indicating that the stimulatory effect of NMDA receptors on substance P release is able to surmount the inhibitory effect of GABAB receptors. In the presence of the GABAB antagonist 2‐hydroxysaclofen (100 μm), but not in its absence, stimulation of the dorsal root at 1 or 10 Hz was able to elicit internalization, which was not inhibited by the NMDA receptor antagonist AP‐5 (50 μm) or the channel blocker MK‐801 (10 μm). Therefore, inhibition of substance P release by GABAB receptors is tonic, and in its absence SP release no longer requires NMDA receptor activation.
Sustained activation of hippocampal pyramidal cells by ‘space clamping’ in a running wheel
Authors: Czurkó András , Hirase Hajime , Csicsvari Jozsef , Buzsáki György
Published: March 2003

In contrast to sensory cortical areas of the brain, the relevant physiological inputs to the hippocampus, leading to selective activation of pyramidal cells, are largely unknown. Pyramidal cells are thought to be phasically activated by spatial cues and a variety of sensory and motor stimuli. Here, we used a behavioural ‘space clamp’ method, which involved the confinement of the actively running animal in a defined position in space (running wheel) and kept sensory inputs constant. Twelve percent of the recorded CA1 pyramidal cells were selectively active while the rat was running in the wheel. Cell firing was specific to the direction of running and disappeared after rotating the recording apparatus. The discharge frequency of pyramidal cells and interneurons was sustained as long as the rat ran continuously in the wheel. Furthermore, the discharge frequency of pyramidal cells and interneurons increased with increasing running velocity, even though the frequency of hippocampal theta waves remained constant. The discharge frequency of some ‘wheel‐related’ pyramidal cells could increase more than 10‐fold between 10 and 100 cm/s, whereas the firing rate of ‘non‐wheel’ cells remained constantly low. We hypothesize that: (i) a necessary condition for place‐specific discharge of hippocampal pyramidal cells is the presence of theta oscillation; and (ii) relevant stimuli can tonically and selectively activate hippocampal pyramidal cells as long as theta activity is present.
Intrathecal galanin alleviates allodynia‐like behaviour in rats after partial peripheral nerve injury
Authors: Hao Jing‐Xia , Shi Tie‐Jun , Xu Isabella S. , Kaupilla Timo , Xu Xiao‐Jun , Hökfelt Tomas , Bartfai Tamas , Wiesenfeld‐Hallin Zsuzsanna
Published: March 2003

We have previously suggested that the neuropeptides galanin and galanin message‐associated peptide (GMAP) may have an inhibitory role in spinal nociception. The present study examined the effects of intrathecal (i.t.) administration of these two peptides on allodynia‐like behaviours in response to mechanical and cold stimulation in rats after photochemically induced ischaemic peripheral nerve injury. I.t. galanin significantly alleviated the mechanical‐ and cold‐allodynia‐like behaviours in nerve injured rats, and was not associated with motor impairment or sedation. I.t. GMAP relieved mechanical allodynia much less than galanin. I.t. M‐35, a high‐affinity galanin receptor antagonist, did not significantly alter the response of the rats to mechanical or cold stimulation. At 1 or 2 weeks postinjury, around 15% of dorsal root ganglion (DRG) neuron profiles showed galanin‐like immunoreactivity. These profiles were mostly small sized. Although the number of galanin positive cells was thus increased in the DRG in the present model, the increase was substantially less than after complete sciatic nerve section, as previously shown.

 The present results showed that spinal administration of galanin inhibited some abnormal pain‐like behaviours in rats after partial peripheral nerve injury. These results further support an inhibitory function for galanin in nociception. However, endogenous galanin may not play a significant role in suppressing nociceptive input after partial ischaemic peripheral nerve injury, as the upregulation of galanin is moderate.
The effects of lesions to the anterior thalamic nuclei on object–place associations in rats
Authors: Sziklas V. , Petrides M.
Published: March 2003

Rats with lesions of the anterior nuclei of the thalamus were trained postoperatively on two spatial conditional associative learning tasks. In the first task, the rats were required to choose one or the other of two objects depending on the location in which they were found. In the second task, the animals learned to turn left or right depending on which one of two visual cues was presented. A third experiment examined the effects of damage to the anterior thalamic nuclei on the eight‐arm radial maze, a spatial working memory task. Damage of the anterior thalamic nuclei impaired performance on the radial maze task and the conditional task requiring associations between objects and their location. By contrast, rats with anterior thalamic lesions were able to acquire, at a rate comparable with that of operated control animals, the conditional task requiring associations between objects and body turns. These findings suggest that lesions to the anterior thalamic nuclei result in a general impairment in learning about allocentric spatial information without disrupting the learning of egocentric spatial information.
Redefining the functional organization of working memory processes within human lateral prefrontal cortex
Authors: Owen Adrian M. , Herrod Nicholas J. , Menon David K. , Clark John C. , Downey Steve P. M. J. , Carpenter T. Adrian , Minhas Pawanjit S. , Turkheimer Federico E. , Williams Emma J. , Robbins Trevor W. , Sahakian Barbara J. , Petrides Michael , Pickard John D.
Published: March 2003

It is widely held that the frontal cortex plays a critical part in certain aspects of spatial and non‐spatial working memory. One unresolved issue is whether there are functionally distinct subdivisions of the lateral frontal cortex that subserve different aspects of working memory. The present study used positron emission tomography (PET) to demonstrate that working memory processes within the human mid‐dorsolateral and mid‐ventrolateral frontal regions are organized according to the type of processing required rather than according to the nature (i.e. spatial or non‐spatial), of the information being processed, as has been widely assumed. Two spatial working memory tasks were used which varied in the extent to which they required different executive processes. During a ‘spatial span’ task that required the subject to hold a sequence of five previously remembered locations in working memory a significant change in blood‐flow was observed in the right mid‐ventrolateral frontal cortex, but not in the anatomically and cytoarchitectonically distinct mid‐dorsolateral frontal‐lobe region. By contrast, during a ‘2‐back’ task that required the subject to continually update and manipulate an ongoing sequence of locations within working memory, significant blood flow increases were observed in both mid‐ventrolateral and mid‐dorsolateral frontal regions. When the two working memory tasks were compared directly, the one that emphasized manipulation of information within working memory yielded significantly greater activity in the right mid‐dorsolateral frontal cortex only. This dissociation provides unambiguous evidence that the mid‐dorsolateral and mid‐ventrolateral frontal cortical areas make distinct functional contributions to spatial working memory and corresponds with a fractionation of working memory processes in psychological terms.
Excitatory synaptic transmission and its modulation by PKC is unchanged in the hippocampus of GAP‐43‐ deficient mice
Authors: Capogna Marco , Fankhauser Christoph , Gagliardini Valeria , Gähwiler Beat H. , Thompson Scott M.
Published: March 2003

We compared excitatory synaptic transmission between hippocampal pyramidal cells in dissociated hippocampal cell cultures and in area CA3 of hippocampal slice cultures derived from wild‐type mice and mice with a genetic deletion of the presynaptic growth associated protein GAP‐43. The basal frequency and amplitude of action potential‐dependent and ‐independent spontaneous excitatory postsynaptic currents were similar in both groups. The probability that any two CA3 pyramidal cells in wild‐type or GAP‐43 knockout (–/–) slice cultures were synaptically connected was assessed with paired recordings and was not different. Furthermore, unitary synaptic responses were similar in the two genotypes. Bath application of phorbol 12,13‐diacetate (0.6–3 μm) elicited a comparable increase in the frequency of miniature excitatory synaptic currents in wild‐type and GAP‐43 (–/–) cultures. This effect was blocked by the protein kinase C inhibitor, bisindolylmaleimide I (1.2 μm). Finally, 3 μm phorbol 12,13‐diacetate potentiated the amplitude of unitary synaptic currents to a comparable extent in wild‐type and GAP‐43 (–/–) slice cultures. We conclude that GAP‐43 is not required for normal excitatory synaptic transmission or the potentiation of presynaptic glutamate release mediated by activation of protein kinase C in the hippocampus.
Depolarization regulates cyclin D1 degradation and neuronal apoptosis: a hypothesis about the role of the ubiquitin/proteasome signalling pathway
Authors: Boutillier Anne‐Laurence , Kienlen‐Campard Pascal , Loeffler Jean‐Philippe
Published: March 2003

Depolarization and subsequent calcium entry exert essential neuroprotective effects but the ultimate effector by which calcium blocks apoptosis is not known. Here we show that inhibition of calcium entry into cerebellar neurons by switching from high to low extracellular K+ concentrations (30–5 mm) induces apoptosis, that correlates with a rapid accumulation of cyclin D1 (CD1), an early marker of the G1/S transition of the cell cycle. These effects on apoptosis and cyclin D1 are mimicked either by blocking calcium entry into neurons (LaCl3, 100 μm or nifedipine, 10–6 m) or by inhibiting the calcium/calmodulin pathway (calmidazolium, 10–7 m). The increased CD1 protein levels do not result from a transcriptional upregulation of the CD1 gene by the Ca2+/calmodulin pathway but rather reflect an accumulation due to the lack of degradation by the proteasome‐dependent pathway. Specific proteasome antagonists: carbobenzoxyl‐leucinyl‐leucinyl‐norvalinal‐H (MG‐115), carbobenzoxyl‐leucinyl‐leucinyl‐leucinal‐H (MG‐132) and clasto‐lactacystin β‐lactone, induce neuronal apoptosis by themselves. Finally, this pathway is functional only at neuroprotective concentrations of K+ (30 mm), suggesting that calcium/CamK signalling pathway may regulate neuronal death by regulating the proteasome‐mediated degradation activity of rapidly turning‐over proteins (constitutively expressed genes or pre‐existing pools of mRNA).
Effect of catecholamines on the hyperpolarization‐activated cationic Ih and the inwardly rectifying potassium IKir currents in the rat substantia nigra pars compacta
Authors: Cathala L. , Paupardin‐Tritsch D.
Published: March 2003

Whole‐cell ruptured‐patch and perforated‐patch recordings were used in principal neurons of the rat substantia nigra pars compacta (SNc) to study the effect of catecholamines both on the hyperpolarization‐activated cationic (Ih) and the inwardly rectifying potassium (IKir) currents. In internal potassium, a 2 min bath application of noradrenaline (NA; 50 μm) or dopamine (DA; 50 μm) both inhibited Ih and induced an outward current associated with an increase in IKir conductance. These two effects recovered poorly after wash‐out. Protein kinase A (PKA), protein kinase C (PKC) and phosphatases 1 and 2A inhibitors did not modify the NA and DA effects on the amplitude of Ih and IKir currents. They also had no effect on the recovery of the catecholamine responses. In perforated‐patch experiments, NA and DA also induced an inhibition of Ih and revealed an outward current associated with an increase in conductance. However, both effects recovered in less than 5 min following the wash‐out. These results indicate that neither PKA, PKC, nor phosphatases 1 or 2A were required in the NA and DA modulation of these two currents and that an intracellular factor, that could be either washed‐out or inversely up‐regulated in the ruptured‐patch configuration, was implicated in the recovery of both effects. In the presence of external barium (300 μm) or internal caesium which both blocked the outward current and the increase in conductance, neither NA nor DA affected Ih, suggesting that the effect on Ih observed is secondary to the activation of the IKir channels. Increasing chloride conductance of the cell by activation of GABAA receptors also induced an inhibition of Ih. All together these results suggest that the NA or DA induced inhibition of Ih could result from an occlusion of Ih by a space‐clamp effect.
Distribution of histamine H3‐receptor binding in the normal human basal ganglia: comparison with Huntington's and Parkinson's disease cases
Authors: Goodchild R. E. , Court J. A. , Hobson I. , Piggott M. A. , Perry R. H. , Ince P. , Jaros E. , Perry E. K.
Published: March 2003

It is now widely recognized that histamine acts as a neurotransmitter in the mammalian central nervous system. Three selective histamine receptors have been described, all of which are present in the basal ganglia. This study is a detailed, quantitative, autoradiographical examination of the densities of histamine H3‐receptors in coronal sections of human basal ganglia, using the selective ligand [3H]‐(R)‐α‐methylhistamine. [3H]‐(R)‐α‐methylhistamine binding was highest within the external and internal segments of the globus pallidus together with the substantia nigra. High levels were also found in the striatum, where density was significantly higher (P < 0.05) at a pre‐, as opposed to post‐, anterior commissure coronal level. Within the striatum, binding was noticeably higher in both the nucleus accumbens and acetylcholinesterase‐deficient striosomes, while being undetectable in the subthalamic nucleus and very low in both the ventroanterior and ventrolateral thalamic nuclei. An intermediate level of binding, often with a laminar distribution, was seen in the insular cortex. [3H]‐(R)‐α‐methylhistamine binding was also examined in both Parkinson's disease and Huntington's disease. No difference from control receptor density was found in any area examined in Parkinson's disease, while values were significantly lower in caudate (P < 0.001), putamen (P < 0.001), external (P < 0.001) and internal (P < 0.05) globus pallidus, although not the insular cortex, in Huntington's disease cases. These data suggest that H3‐receptors are present upon striatonigral projection neurons of the direct and indirect movement pathways thus providing histaminergic control over the activity of both these circuits.
The protein kinase C (PKC) substrate GAP‐43 is already expressed in neural precursor cells, colocalizes with PKCη and binds calmodulin
Authors: Esdar Christina , Oehrlein Silke A. , Reinhardt Sigrid , Maelicke Alfred , Herget Thomas
Published: March 2003

Expression of the growth‐associated protein of 43‐kDa (GAP‐43), which is described as a postmitotic, neuron‐specific major protein kinase C (PKC) substrate, was investigated in the murine embryonic carcinoma cell line PCC7‐Mz1 which develops into a brain‐tissue‐like pattern of neuronal, fibroblast‐like and astroglial cells upon stimulation with all‐trans retinoic acid (RA). GAP‐43 expression was very low in stem cells, but increased on mRNA and protein level within the 12 h after differentiation was initiated. While the P1 promoter of the GAP‐43 gene gave rise to a 1.6‐kb mRNA and was already active at a very low level in PCC7‐Mz1 stem cells, transcription of the P2 promoter, which resulted in a 1.4‐kb mRNA, was completely blocked in stem cells but increased rapidly after RA treatment.

Within the first 2 days of neural differentiation, GAP‐43 was localized with the cytoplasmic membrane and the Golgi complex of proliferating neural precursor cells. Then, GAP‐43 was translocated to the growth cones and neurites, and from day 6, when neurons began to acquire polarity, the protein was found in the axons. GAP‐43 was never detected in the non‐neuronal PCC7‐Mz1 derivatives, i.e. in fibroblasts or glial cells.

In the foetal rat brain (prenatal day F11), GAP‐43 was expressed in the optic stalk, the lense plakode and in the postmitotic neurons of the marginal zone of the hindbrain. Moreover, in a layer between the ventricular and marginal zone of the hindbrain (F13) and forebrain (F15), GAP‐43 was already expressed in mitotic neural precursor cells.

In PCC7‐Mz1 cultures, 2 days after addition of RA, GAP‐43 became phosphorylated upon activation of PKC, and colocalized specifically with the novel PKC isoform η. Phosphorylation of GAP‐43 caused a disruption of its complex with calmodulin. These data demonstrate that GAP‐43 is already a functional PKC substrate in prolific neuronal precursor cells, and may participate in neuronal cell lineage determination.
Proteolytic processing, axonal transport and differential distribution of chromogranins A and B, and secretogranin II (secretoneurin) in rat sciatic nerve and spinal cord
Authors: Li Jia‐Yi , Leitner Bernd , Lovisetti‐Scamihorn Paola , Winkler Hans , Dahlström Annica
Published: March 2003

The chromogranin family comprises chromogranin A and B, and secretogranin II. The present study has focused on the axonal transport of chromogranins/secretogranin II and their detailed distribution in peripheral nerves and the spinal cord. With radioimmunoassay (RIA) and column chromatography, we first studied the processing of chromogranin B and secretogranin II during axonal transport. No larger precursors of these peptides were detected in the sciatic nerves, indicating that they are already processed to a high degree early during axonal transport. We also analysed nerve segments above and below a crush, using RIA, in order to compare these accumulation data with those obtained by the cytofluorimetric‐scanning (CFS) technique. For the latter technique, the amounts of accumulation distal to the crush (presumably representing recycling and retrogradely transported peptides) were 30–40% of the amounts in the proximal accumulation for chromogranin A and secretoneurin, in contrast to chromogranin B, which showed 15% recycling. With the RIA, the corresponding values for secretoneurin and PE‐11 (antibody against chromogranin B) were 42% and 14%, respectively. Therefore, the data obtained by CFS were in excellent agreement with those obtained by RIA. In crushed sciatic nerves, chromogranin A was present in large axons as well as in small‐ and medium‐sized axons. Chromogranin B was mainly restricted to large axons, while secretoneurin was localized to bundles of small axons. This differential distribution was also found in the spinal roots and in the peripheral terminals. Chromogranin A was present in both ventral and dorsal roots, and chromogranin B was detected in ventral roots and in large sensory axons in the dorsal roots. Secretoneurin was dominant in the dorsal root. Double‐labelling studies with antibodies against choline acetyltransferase/vesicular acetylcholine transporter, or against tyrosine hydroxylase, confirmed that chromogranin A was distributed in cholinergic, sensory, as well as adrenergic neurons. Chromogranin B was mainly present in cholinergic motor neurons and large sensory neurons, and secretoneurin was restricted to adrenergic and sensory neurons. The present study demonstrates that chromogranins A and B, and secretoneurin are transported with fast axonal transport in the peripheral nerves, with different amounts of recycling, and that they are differentially distributed in different types of neurons in the peripheral nervous system and the spinal cord, suggesting that each of them may play a special role in subsets of neurons.
Heparan sulphate and HB‐GAM (heparin‐binding rowth‐associated molecule) in the development of the thalamocortical pathway of rat brain
Authors: Kinnunen Anu , Niemi Maria , Kinnunen Tarja , Kaksonen Marko , Nolo Riitta , Rauvala Heikki
Published: March 2003

Extracellular matrix (ECM) molecules, such as laminin, tenascin, chondroitin sulphate proteoglycans and heparan sulphate proteoglycans have been suggested to have ‘signpost’ and directing roles in the formation of axonal projections in cortical development. We show here that the expression of the neurite outgrowth‐promoting protein heparin‐binding growth‐associated molecule (HB‐GAM) and N‐syndecan, a transmembrane heparan sulphate proteoglycan previously isolated as a receptor for HB‐GAM, is spatiotemporally associated with the developing thalamocortical pathway in the rat brain. Using in situ hybridization, thalamic neurons were shown to express mRNA for N‐syndecan, and in vitro, thalamic neurons grew more neurites on HB‐GAM than on laminin. The HB‐GAM‐induced neurite outgrowth in thalamic neurons was inhibited by heparitinase, heparin, soluble N‐syndecan and by an excess of soluble HB‐GAM in the culture medium. In a pathway assay, thalamic neurons selectively preferred attaching and growing neurites on matrices containing HB‐GAM than on those containing fibronectin or laminin alone, suggesting that HB‐GAM may modulate the effect of other ECM proteins. On an unfixed brain slice preparation, thalamic neurons repeatedly showed a typical neurite outgrowth and attachment pattern resembling the expression pattern of HB‐GAM. On the brain slices, the neurite outgrowth was significantly inhibited by heparitinase, heparin and soluble HB‐GAM, thus displaying features of neurite outgrowth on matrix‐bound HB‐GAM. Our results suggest that HB‐GAM is important for the neurite outgrowth of thalamic neurons and it may function as an ECM‐bound guidance cue for thalamic neurons that possess HB‐GAM‐binding heparan sulphates on their cell membrane.
Stability and plasticity of primary afferent projections following nerve regeneration and central degeneration
Authors: Belyantseva Inna A. , Lewin Gary R.
Published: March 2003

Sensory neurons of the dorsal root ganglia (DRG) regenerate their peripheral axons with relative ease following a nerve lesion. The capacity for central regeneration appears more limited. However, after nerve lesion, some DRG neurons gain a regenerative advantage to sprout centrally. We developed a lesion model in the rat to test whether, after prior lesion of their peripheral axons, subsets of cutaneous afferents benefit differently in their ability to sprout into adjacent spinal segments denervated by dorsal rhizotomy. We found that under identical circumstances, myelinated sensory neurons, small‐diameter peptidergic sensory neurons containing calcitonin gene related peptide (CGRP), and small‐diameter nonpeptidergic neurons that bind the lectin from the plant Griffonia simplificolia, isolectin B4 (IB4) differ dramatically in their ability to regenerate centrally. Myelinated afferent terminals labelled transganglionically with cholera‐toxin β‐subunit gain a small advantage in collaterally sprouting into the adjacent denervated neuropil in lamina III after prior peripheral nerve lesion. This central regenerative response was not mimicked by experimentally induced inflammation of sensory neuron cell bodies. Intact and unlesioned sensory neurons positive for CGRP sprout vigorously into segments denervated by rhizotomy in a nonsomatotopic manner. In contrast, IB4‐positive sensory neurons maintain a somatotopic distribution centrally, which is not altered by prior nerve lesion. These data reveal a remarkably heterogeneous response to regeneration‐promoting stimuli amongst three different types of cutaneous sensory neurons. In particular, the divergent responses of peptidergic and nonpeptidergic sensory neurons suggests profound functional differences between these neurochemically distinct neurons.
Expression of unphosphorylated class III β‐tubulin isotype in neuroepithelial cells demonstrates neuroblast commitment and differentiation
Authors: Fanarraga M. L. , Avila J. , Zabala J. C.
Published: March 2003

Neuronal microtubules have unique stability properties achieved through developmental regulation at the expression and post‐translational levels on tubulins and microtubule associated proteins. One of the most specialized tubulins specific for neurons is class‐III β‐tubulin (also known as β6‐tubulin). Both the upregulation and the post‐translational processing of class‐III β‐tubulin are believed to be essential throughout neuronal differentiation. The present investigation documents the temporal and spatial patterns of class‐III β‐tubulin expression throughout neurogenesis. For this study a novel polyclonal antiserum named U‐β6, specific to unphosphorylated class‐III β‐tubulin has been developed, characterized and compared with its commercial homologue TuJ‐1. Our experiments indicate that the two antibodies recognize different forms of class‐III β‐tubulin both in vitro and in vivo. Biochemical data revealed that U‐β6 bound unphosphorylated soluble class‐III β‐tubulin specifically, while TuJ‐1 recognized both the phosphorylated and unphosphorylated forms of the denatured protein. In vivo U‐β6 was associated with neurogenesis and labelled newly committed CNS and PNS neuroblasts expressing neuroepithelial cytoskeletal (nestin and vimentin) and surface markers (the anti‐ganglioside supernatant, A2B5 and the polysialic acid neural adhesion molecule, PSA‐NCAM), as well as differentiating neurons. These studies with U‐β6 illustrate three main developmental steps in the neuronal lineage: the commitment of neuroepithelial cells to the lineage (U‐β6 +ve/TuJ‐1 –ve cells); a differentiation stage (U‐β6 +ve/TuJ‐1 +ve cells); and, finally, neuronal maturation correlating with a drop in unphosphorylated class‐III β‐tubulin immunostaining levels. These investigations also conclude that U‐β6 is an earlier marker than TuJ‐1 for the neuronal lineage in vivo, and it is thus the earliest neuronal lineage marker known so far.
Specific expression of ezrin, a cytoskeletal‐membrane linker protein, in a subset of chick retinotectal and sensory projections
Authors: Takahashi Masakazu , Yamagata Masahito , Noda Masaharu
Published: March 2003

Lamina‐specific neuronal connections are a fundamental feature in many parts of the vertebrate central nervous system. In the chick, the optic tectum is the primary visual centre, and it has a multilaminated structure consisting of 15 laminae, of which only three or four receive retinal projections. Each of the retinorecipient laminae establishes synaptic connections selectively from one of a few subsets of retinal ganglion cells (RGCs). We have generated a series of monoclonal antibodies that appear to stain only one of the retinorecipient laminae. One of these, TB4, stained lamina F which receives inputs from a subpopulation of ≈ 10–20% of RGCs which express the presynaptic acetylcholine receptor β2‐subunit. TB4 recognized a single 79‐kDa protein on immunoblotting. cDNA cloning and immunochemical analysis revealed that the TB4 antigen molecule was ezrin, a cytoskeletal‐membrane linker molecule belonging to the ezrin‐radixin‐moesin family. Unilateral enucleation of the eye, both prior to and after the establishment of retinotectal projections, attenuated the lamina‐selective staining with TB4 in the contralateral tectum, suggesting that ezrin is anterogradely transported from RGCs to lamina F. Ezrin was thus expressed in a subset of RGCs that project to lamina F. Similar subset‐selective expression and resultant lamina‐selective distribution of ezrin were also observed in the lamina‐specific central projections from the dorsal root ganglia. The staining pattern with TB4 in the dorsal root ganglia and spinal cord indicated that high expression of ezrin was restricted in cutaneous sensory neurons, but not in muscle sensory neurons. Since ezrin modulates cell morphology and cell adhesion profiles by linking membrane proteins with the cytoskeleton, it was suggested that ezrin is involved in the formation and/or maintenance of lamina‐specific connections for neuronal subpopulations in the visual and somatosensory systems.
Do superior colliculus projection zones in the inferior pulvinar project to MT in primates?
Authors: Stepniewska Iwona , Qi Hui‐Xin , Kaas Jon H.
Published: March 2003

In order to determine the relationship of superior colliculus inputs to thalamic neurons projecting to the middle temporal visual area (MT), injections of wheat germ agglutinin conjugated with horseradish peroxidase were placed in the superior colliculus of three owl monkeys, with injections of Fast Blue in the MT. The locations of labelled terminals and neurons in the posterior thalamus were related to four architectonically distinct nuclei of the inferior pulvinar (Stepniewska & Kaas, Vis. Neurosci.14, pp.1043–1060, 1997). Fast Blue injections in the MT labelled neurons largely in the medial nucleus of the inferior pulvinar. A few labelled neurons were found in the adjoining central medial nucleus of the inferior pulvinar, as well as in the lateral pulvinar and the dorsal lateral geniculate nucleus. Superior colliculus inputs were most dense in the posterior and medial nuclei of the inferior pulvinar. There were sparser inputs to the central lateral nucleus of the inferior pulvinar, locations in the lateral and medial pulvinar, and the dorsal lateral geniculate nucleus. The results indicate that the medial nucleus of the inferior pulvinar, the major projection zone to the MT, does not receive a significant input from the superior colliculus.
Chronic morphine exposure and spontaneous withdrawal are associated with modifications of dopamine receptor and neuropeptide gene expression in the rat striatum
Authors: Georges François , Stinus Luis , Bloch Bertrand , Moine Catherine Le
Published: March 2003

The influence of chronic morphine and spontaneous withdrawal on the expression of dopamine receptors and neuropeptide genes in the rat striatum was investigated. Morphine dependence was induced by subcutaneous implantation of two morphine pellets for 6 days. Rats were made abstinent by removal of the pellets 1, 2 or 3 days before they were killed. The mRNA levels coding for D1‐ and D2‐dopamine receptors, dynorphin, preproenkephalin A and substance P were determined by quantitative in situ hybridization. The caudate putamen and the nucleus accumbens showed equivalent modifications in dopamine receptor and neuropeptide gene expression. After 6 days of morphine, a decrease in D2‐dopamine receptor and neuropeptide mRNA levels was observed (– 30%), but there was no change in D1‐dopamine receptor mRNA. In abstinent rats, both D1‐ and D2‐dopamine receptor mRNA levels were decreased 1 day after withdrawal (– 30% compared with chronic morphine). In contrast, neuropeptide mRNA levels were unaffected when compared with those observed after 6 days of morphine. During the second and third day of withdrawal, there was a gradual return to the levels seen in the placebo‐treated group, for both dopamine receptor and neuropeptide mRNAs. Phenotypical characterization of striatal neurons expressing μ and κ opioid receptor mRNAs showed that, in striatonigral neurons, both mRNAs were colocalized with D1‐receptor and Dyn mRNAs. Our results suggest that during morphine dependence, dopamine and morphine exert opposite effects on striatonigral neurons, and that effects occurring on striatopallidal neurons are under dopaminergic control. We also show that withdrawal is associated with a down regulation of the postsynaptic D1 and D2 receptors.
Distinct populations of hypothalamic dopaminergic neurons exhibit differential responses to brain‐derived neurotrophic factor (BDNF) and neurotrophin‐3 (NT3)
Authors: Loudes C. , Petit F. , Kordon C. , Faivre‐Bauman A.
Published: March 2003

We have previously demonstrated that differentiation of hypothalamic dopaminergic (DA) neurons can be induced in culture by their pituitary intermediate lobe target cells, through both membrane and diffusible factors. We also showed that subpopulations of DA neurons from the arcuate nucleus only, not the periventricular area, can respond to the target. Here we investigated the possibility that both neuronal subsets could also respond differentially to brain‐derived neurotrophic factor (BDNF) or neurotrophin‐3 (NT3). Addition of NT3, but not BDNF, enhanced growth and branching of neurites, tyrosine hydroxylase (TH) as well as increasing levels of cultured arcuate DA neurons. Conversely, BDNF, but not NT3, affected the same parameters in cultured periventricular DA neurons. The neurotrophins thus affect DA neurons in a structure and neuronal type‐selective manner, since general neuronal markers were not affected by either neurotrophin. Neurotrophin effects were reversed by addition of specific antibodies directed against them or their respective receptors, TrkB or TrkC. By themselves, the antibodies inhibited development of DA neurons below that of control cultures, suggesting involvement of endogenous neurotrophins. BDNF and NT3 were indeed found in both arcuate and periventricular neurons and in the intermediate lobe. BDNF was always present as the mature peptide. The mature form of NT3 was only detected in the periventricular area; a precursor‐like heavier form was present in all tissues studied. The present data suggest that NT3, but not BDNF, could participate in the differentiating action of intermediate lobe cells on arcuate DA neurons.
Role of α2C‐adrenoceptor subtype in spatial working memory as revealed by mice with targeted disruption of the α2C‐adrenoceptor gene
Authors: Tanila H. , Mustonen K. , Sallinen J. , Scheinin M. , Riekkinen P.
Published: March 2003

The role of the α2C‐adrenoceptor subtype in mediating the beneficial effect of α2‐adrenoceptor agonists on spatial working memory was studied in adult mice with targeted inactivation of the α2C‐receptor gene (KO) and their wild‐type controls (WT). A delayed alternation task was run in a T‐maze with mixed delays varying from 20 s to 120 s. Dexmedetomidine, a specific but subtype non‐selective α2‐adrenoceptor agonist, dose‐dependently decreased the total number of errors. The effect was strongest at the dose of 5 μg/kg (s.c.), and was observed similarly in KO and WT mice. KO mice performed inferior to WT mice due to a higher number of perseverative errors. Dexmedetomidine slowed initiation of the motor response in the start phase at lower doses in WT mice than in KO mice but no such difference was observed in the return phase of the task, suggesting involvement of α2C‐adrenoceptors in the cognitive aspect of response preparation or in response sequence initiation. According to these findings, enhancement of spatial working memory is best achieved with α2‐adrenoceptor agonists which have neither agonistic nor antagonistic effects at the α2C‐adrenoceptor subtype.
Inducing placebo respiratory depressant responses in humans via opioid receptors
Authors: Benedetti Fabrizio , Amanzio Martina , Baldi Sergio , Casadio Caterina , Maggi Giuliano
Published: March 2003

Several lines of evidence indicate that placebos produce analgesia through the activation of endogenous opioid systems. Recently, we showed that placebos may also produce respiratory depressant responses, a typical side‐effect of narcotics, when a subject had a prior experience of respiratory depression in the course of narcotic treatment. In the present study, we report that the placebo respiratory depression can be induced after repeated administrations of the partial opioid agonist buprenorphine. The placebo respiratory depressant effect that resulted from the buprenorphine conditioning was completely blocked by a dose of 10 mg of naloxone, indicating that it was mediated by endogenous opioids. These findings show that placebos act, via the activation of opioid receptors, not only on pain mechanisms but on the respiratory centres as well, thus mimicking a typical side‐effect of narcotics. In addition, the experimental procedure we used did not produce any expectation of respiratory depression and, similarly, the subjects did not notice any sign of respiratory discomfort. Thus, the placebo respiratory depression elicited in the present study cannot be explained on the basis of cognitive or motivational mechanisms. Rather, it appears to be a sequence effect due to learning, thus suggesting a conditioning mechanism mediated by endogenous opioids.
Inhibition of collagen IV deposition promotes regeneration of injured CNS axons
Authors: Stichel Christine C. , Hermanns Susanne , Luhmann Heiko J. , Lausberg Friederike , Niermann Heike , D'Urso Donatella , Servos Gisela , Hartwig Hans‐Georg , Müller Hans Werner
Published: March 2003

Scarring impedes axon regrowth across the lesion site and is one major extrinsic constraint to effective regeneration in the adult mammalian central nervous system. In the present study we determined whether specific biochemical or immunochemical modulation of one major component of the scar, the basal membrane (BM), would provide a means to stimulate axon regeneration in the mechanically transected postcommissural fornix of the adult rat. Basal membrane developed within the first 2 weeks after transection in spatiotemporal coincidence with the abrupt growth arrest of spontaneously regrowing axons. Local injection of anticollagen IV antibodies or α, α′‐dipyridyl, an inhibitor of collagen triple helix formation and synthesis, significantly reduced lesion‐induced BM deposition. This treatment allowed massive axon elongation across the lesion site. Anterograde tracing provided unequivocal evidence that regenerating axons follow their original pathway, reinnervate the appropriate target, the mammillary body, and become remyelinated with compact myelin. Presynaptic electrophysiological recordings of regenerated fibre tracts showed recovery to nearly normal conduction properties. Our results indicate that lesion‐induced BM is an impediment for successful axonal regeneration and its reduction is a prerequisite and sufficient condition for regrowing axons to cross the lesion site.
Brain location and visual topography of cortical area V6A in the macaque monkey
Authors: Galletti Claudio , Fattori Patrizia , Kutz Dieter F. , Gamberini Michela
Published: March 2003

The brain location, extent and functional organization of the cortical visual area V6A was investigated in macaque monkeys by using single cell recording techniques in awake, behaving animals. Six hemispheres of four animals were studied. Area V6A occupies a horseshoe‐like region of cortex in the caudalmost part of the superior parietal lobule. It extends from the medial surface of the brain, through the anterior bank of the parieto‐occipital sulcus, up to the most lateral part of the fundus of the same sulcus. Area V6A borders on areas V6 ventrally, PEc dorsally, PGm medially and MIP laterally. Of 1348 neurons recorded in V6A, 61% were visual and 39% non‐visual in nature. The visual neurons were particularly sensitive to orientation and direction of movement of visual stimuli. The inferior contralateral quadrant was the most represented one. Visual receptive fields were also found in the inferior ipsilateral quadrant and in the upper visual field. Receptive fields were on average smaller in the lower visual field than in the upper one. Both central and peripheral parts of the visual field were represented. Large parts of the visual field were represented in small regions of area V6A, and the same regions of the visual field were re‐represented many times in different parts of this area, without any apparent topographical order. The only reliable sign of retinotopic organization was the predominance of central representation dorsally and far periphery ventrally. The functional organization of area V6A is discussed in the view that this area could be involved in the control of reaching out and grasping objects.
Effects of dorsolateral spinal lesions on stretch reflex threshold and stiffness in awake cats
Authors: Taylor Julian , Munson John , Vierck Jr Charles
Published: March 2003

Measurements of threshold angle and incremental dynamic stiffness (IDS) were derived from triceps surae stretch reflexes, elicited by ramp and hold flexion at the ankle joint of four cats that were tested while awake. Stretch reflexes were assessed from trials that began from different ankle joint start positions or were matched using a post‐hoc analysis for initial background force during testing sessions before and following unilateral lesions of the dorsolateral funiculus at levels ranging from T13 to L3. Unilateral lesions of the dorsolateral funiculus (DLF) produced significant ipsilateral decreases in stretch reflex threshold and increases in reflex gain, measured as incremental dynamic stiffness (IDS). ancova testing indicated that the reduction in threshold, but not the increase in IDS, was dependent upon the level of background force. Reflex testing from different start angles demonstrated that DLF lesions diminished the correlation between threshold and IDS. Intravenous infusion of ketamine dose‐dependently reduced IDS, compared with testing in the unanaesthetized state. Postoperative reflex testing during infusion of ketamine at 22.2 mg/kg per h, when electromyographic responses were reduced to 24% of control levels, abolished differences in IDS between the ipsilateral and contralateral hindlimbs. These and related observations suggest that the postoperative increase in IDS in awake animals was not due to an increase in passive stiffness.
Neurotrophic factors modulate hair cells and their potassium currents in chick otocyst explants
Authors: Sokolowski Bernd H. A. , Csus Jolan , Hafez Osama I. , Haggerty Holly S.
Published: March 2003

Neurotrophins, retinoids and their receptors are present in the sensory epithelia of the inner ear during development. We show that these factors modulate the proliferation of hair cells and their K+‐currents when the embryonic day 3 (ED 3) presumptive inner ear (i.e. otocyst) is maintained in organ culture. All trans‐retinoic acid (RA) increases hair cell differentiation and enhances the acquisition of outward currents, including a delayed rectifier and a fast activating, transient type, voltage‐gated potassium current. In contrast, brain‐derived neurotrophic factor (BDNF) and neurotrophin‐3 (NT‐3) decrease ionic current activity, and the addition of RA with the neurotrophins enhances this inhibitory response in an age‐dependent manner. We measured the total number of cells per explant over time to determine precisely when and how these factors inhibit explant growth. We found that high concentrations of BDNF and NT‐3 administered together, and low concentrations of both neurotrophins combined and administered with RA suppress otocyst cell numbers after 24 h in vitro. This suppressive response is induced by RA and NT‐3, not by RA and BDNF. The suppressive or inhibitory influence of NT‐3 and RA is the result of NT‐3 binding to the low affinity receptor, p75NTR, not the result of RA increasing mRNA levels for the high affinity receptor, trkC. However, trk may act with p75NTR, as disruption of trk signalling alleviates the inhibitory response induced by NT‐3 and RA. Our data suggest that various combinations and/or concentration gradients of these factors can differentially regulate inner ear development and hair cell excitability.