bluuebunny/crossref_metadata_2025 huggingface dataset:
Long‐Term Depression in Rat Cerebellum Requires both NO Synthase and NO‐sensitive Guanylyl Cyclase
Authors: Boxall Andrew R. , Garthwaite John
Published: April 2006

Long‐term depression (LTD) of synaptic transmission between parallel fibres and Purkinje cells is a well‐known example of synaptic plasticity taking place in the cerebellum. Nitric oxide (NO) has been implicated in synaptic plasticity in other brain areas, but its function in cerebellar LTD is controversial. Even when an involvement is suggested, the NO signal transduction pathway is unclear. One candidate is the cyclic GMP‐synthesizing enzyme, soluble guanylyl cyclase, whose activity in the brain and elsewhere is powerfully stimulated by NO. By recording intracellularly from Purkinje cells in cerebellar slices, we demonstrate that blockade of NO synthase completely inhibits LTD induced by pairing parallel fibre stimulation with postsynaptic Ca2+ spike firing. LTD was also blocked by intracellular application of 1H‐[1, 2, 4]oxadiazolo[4, 3‐a]quinoxalin‐1‐one, a recently identified potent and selective inhibitor of soluble guanylyl cyclase. These findings indicate that soluble guanylyl cyclase is required for cerebellar LTD and suggest that this enzyme, located within Purkinje cells, transduces the NO signal in this form of synaptic plasticity.
Leukaemia Inhibitory Factor Induced in the Sciatic Nerve after Axotomy is Involved in the Induction of Galanin in Sensory Neurons
Authors: Sun Y. , Zigmond R. E.
Published: April 2006

Dramatic changes occur in neuropeptide expression in sensory and sympathetic neurons following axonal injury. Based on the finding that the cytokine leukemia inhibitory factor (LIF) plays an important role in mediating these changes in sympathetic neurons, its participation in triggering changes in sensory neurons was examined. By the use of transgenic mice in which the LIF gene had been knocked out, LIF was found to contribute to the induction of galanin expression in dorsal root ganglia (DRG) after sciatic nerve lesion. On the other hand, two other neuropeptide changes that occur in DRG under these conditions, the reduction of substance P and induction of neuropeptide Y, were independent of LIF expression. In the sympathetic superior cervical ganglion, transection of the postganglionic nerves close to the ganglion resulted in a rapid induction of LIF mRNA in the ganglion and in the lesioned nerve trunk. In contrast, transection of the sciatic nerve close to or distant from the DRG caused a rapid induction of LIF mRNA in the lesioned nerve, but not in the DRG. DRG were capable of making substantial amounts of LIF mRNA when placed in explant cultures, but,in vivo, only a slight induction was found even when both central and peripheral nerve processes of these sensory neurons were transected. These latter observations suggest that, in contrast to the superior cervical ganglia, the DRG environment inhibits the lesion‐induced expression of LIFin vivo, and/or explanted DRG produce stimulatory signals not foundin vivo., Together with the data on the induction of galanin, these observations provide evidence that LIF, generated at a site at some distance from the ganglion, is involved in triggering part of the cell body reaction in sensory neurons.
Functional Demarcation of a Border Between Areas V6 and V6A in the Superior Parietal Gyrus of the Macaque Monkey
Authors: Galletti C. , Fattori P. , Battaglini P. P. , Shipp S. , Zeki S.
Published: April 2006

We have compared physiological data recorded from three alert macaque monkeys with separate observations of local connectivity, to locate and characterize the functional border between two related but distinct visual areas on the caudal face of the superior parietal gyrus. We refer to these areas as V6 and V6A. They occupy almost the entire extent of the anterior bank of the parieto‐occipital sulcus, V6A being the more dorsal. These two areas are strongly interconnected. Anatomically, we have defined the border as the point at which labelled axon terminals first adopt a recognizably ‘descending’ pattern in their laminar characteristics, after injections of wheatgerm agglutinin‐horseradish peroxidase into the dorsal half of the gyrus (in presumptive V6A). A similar principle was used to recognize the same border by the pattern of input from area V5, except that in this case the relevant transition in laminar characteristics is that between an ‘intermediate’ pattern (in V6) and an ‘ascending’ pattern (in V6A). V6A was found to be distinct from V6 in a number of its physiological properties. Unlike V6, it contains visually unresponsive cells as well as units with craniotopic receptive fields (‘real‐position’ cells), units tuned to very slow stimulus speeds, units with complex visual selectivities and units with activity related to attention. V6A was also found to have a larger mean receptive field size and scatter than V6. By contrast, response properties related to the basic orientation and direction of moving bar stimuli were indistinguishable between V6 and V6A, as was the influence of gaze direction on cell activity in the two areas. Two‐dimensional maps of the recording sites allowed reconstruction of the V6/V6A border. For comparison, the anatomical results were rendered on two‐dimensional maps of identical format to those used to summarize the physiological data. After normalizing for relative size, the physiological and connectional estimates of the border between V6 and V6A were found to coincide, at least within the range of individual variation between hemispheres. An architectonic map in the same format was also made from a hemisphere stained for myelin and Nissl substance. Area PO, defined by its general density of myelination was not distinct in this material, but several architectural features were traceable and one of these was also found to approximate the V6/V6A border. The particular criteria that distinguish V6 from V6A differ from a recent description of areas PO and POd in the Cebus monkey; we believe it most likely that PO and POd together may correspond to V6.
Differential Distribution of MAP1a and Aldolase c in Adult Mouse Cerebellum
Authors: Touri F. , Hawkes R. , Riederer B. M.
Published: April 2006

MAP1a is a microtubule‐associated protein with an apparent molecular weight of 360 kDa that is found in the axonal and dendritic processes of neurons. Two monoclonal anti‐MAP1a antibodies, anti‐A and anti‐BW6, revealed different epitope distributions in the adult mouse cerebellum. Anti‐A stained Purkinje and granule cells uniformly throughout the cerebellum. In contrast, anti‐BW6 selectively stained the dendrites of a subset of Purkinje cells, revealing parasagittal bands of immunoreactivity in the molecular layer. The compartmentation of the BW6 epitope was compared to the Purkinje cells as revealed by immunostaining with anti‐zebrin II, a well known antigen expressed selectively by bands of Purkinje cells. The anti‐BW6 staining pattern was complementary to the zebrin II bands, the zebrin II‐ Purkinje cells having BW6+ dendrites. These results demonstrate that MAP1a is present in two forms in the mouse cerebellum, one of which is segregated into parasagittal bands. This may indicate a unique MAP1a isoform or may reflect differences in the metabolic states of Purkinje cell classes, and regional differences in their functions.
Characterization of Excitatory Amino Acid Neurotoxicity in N‐methyl‐D‐aspartate Receptor‐deficient Mouse Cortical Neuronal Cells
Authors: Tokita Yoshihito , Bessho Yasumasa , Masu Masayuki , Nakamura Kenji , Nakao Kazuki , Katsuki Motoya , Nakanishi Shigetada
Published: April 2006

Roles and mechanisms of N‐methyl‐D‐aspartate (NMDA) receptors in glutamate neurotoxicity were investigated in cultures of NMDA receptor‐deficient cortical neuronal cells. Mutant mice lacking a functional NMDA receptor were generated by gene targeting of the NR1 NMDA receptor subunit. Cortical neuronal cells prepared from wild‐type NR1+/+, heterozygous NR1+/‐ and homozygous mutant NR1‐/‐ mice at 15–17 days of gestation grew indistinguishably from each other. Brief exposures (5 min) of both NR1+/+ and NR1+/‐ neuronal cells to glutamate or NMDA, but not kainate or α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionate (AMPA), resulted in widespread neuronal degeneration by the following day. In contrast, neither glutamate nor NMDA treatment caused neuronal degeneration in NR1‐/‐ cells, indicating that NMDA receptors are responsible for rapidly triggered glutamate neurotoxicity. The above four compounds were all effectivein inducing the death of NR1+/+ and NR1+/‐ neuronal cells after prolonged exposure (20–24 h). However, NMDA had no neurotoxic effects on NR1‐/‐ cells, although the other three compounds were neurotoxic with potencies comparable to those for NR1+/+ and NR1+/‐ cells. The AMPA and kainate receptors are thus sufficient for inducing slowly triggered glutamate neurotoxicity. Brief exposure of a mixed population of NR1+/+ and NR1‐/‐ neuronal cells to NMDA selectively killed the NMDA receptor‐expressing cells without any appreciable effects on neighbouring NMDA receptor‐deficient cells. This finding further supports a direct and indispensable role for NMDA receptors in NMDA‐evoked neuronal cell death.
Immunocytochemical Evidence that Glutamate is a Neurotransmitter in the Cochlear Nerve: A Quantitative Study in the Guinea‐pig Anteroventral Cochlear Nucleus
Authors: Hackney Carole M. , Osen Kirsten K. , Ottersen Ole Petter , Storm‐Mathisen Jon , Manjaly George
Published: April 2006

The large so‐called type I afferents of the cochlear nerve carry the majority of the auditory input from the cochlea to the cochlear nuclei in the brainstem. These fibres are excitatory and previous studies have suggested they may use glutamate as their neurotransmitter. In the present investigation therefore, antibodies to glutamate and to the glutamate precursor, glutamine, were applied to resin sections of perfusion‐fixed brains and of in vitro brain slices subjected to depolarizing levels of potassium before fixation to study glutamate handling and synaptic release. Ultrathin sections were labelled by the immunogold technique, and the immunoreactivity was quantified by recording the density of gold particles over the various tissue profiles. Non‐primary, presumably inhibitory, terminals and glial processes were used as reference structures. The cochlear primary terminals proved to be strongly immunoreactive for glutamate. The density of glutamate labelling was higher in primary terminals than in non‐primary ones, and lowest in glial processes. The ratio between the mean glutamate and glutamine labelling densities was also higher in primary terminals than in non‐primary ones, and lowest in glial processes in each case. In the primary terminals, the glutamate immunoreactivity was higher over vesicle‐containing regions than over vesicle‐free regions, whilst glutamine was evenly distributed throughout. The in vitro brain slices showed a potassium‐induced, partly calcium‐dependent depletion of glutamate from the primary terminals but not from the non‐primary ones. These observations strongly support the conclusion that glutamate is a neurotransmitter of type I cochlear afferents.
The Effects of Lesions to the Mammillary Region and the Hippocampus on Conditional Associative Learning by Rats
Authors: Sziklas V. , Petrides M. , Leri F.
Published: April 2006

Rats with extensive lesions to the mammillary body region, the hippocampus, or rats which had received a control operation were trained postoperatively on two visuo‐spatial conditional associative learning tasks in which they had to learn to associate spatial cues with particular visual/auditory stimuli. The animals were subsequently trained on a spatial working memory task, the eight‐arm radial maze. Rats with lesions to the mammillary body region were able to acquire the conditional associative learning tasks at a rate comparable to that of operated control animals, whereas those with hippocampal lesions were not. By contrast, rats with a lesion of the mammillary body region or the hippocampus were significantly impaired in comparison with the operated control animals in the radial maze. The findings suggest that lesions to the mammillary body region impair spatial working memory without affecting the capacity to associate particular exteroceptive cues with spatial locations.
Glutamate Depresses Release by Activating Non‐conventional Glutamate Receptors at Crayfish Nerve Terminals
Authors: Parnas I. , Dudel J. , Parnas H. , Ravin R.
Published: April 2006

The present study shows that release of glutamate from crayfish nerve terminals is inhibited at low depolarizing current pulses by glutamate, N‐methyl‐D‐aspartate (NMDA) and quisqualate. These agonists elicit inhibitory effects at concentrations as low as 10‐8 M (quisqualate) and 10‐7 M (glutamate and NMDA). The NMDA‐mediated inhibition is blocked by (±)‐2‐amino‐5‐phosphonovaleric acid (APV). The quisqualate‐mediated inhibition is blocked by 6‐cyano‐7‐nitroquinoxaline‐2, 3‐dione (CNQX). Both CNQX and APV are needed to block glutamate‐mediated inhibition. The inhibition of release is not accompanied by a detectable change in presynaptic membrane conductance at the secondary branch. Using fura‐2, Ca2+ accumulation during repetitive stimulation (100 Hz) was monitored in single release boutons. Inhibition of release, elicited by 10‐4 M glutamate, was not associated with a reduction in the accumulation of Ca2+. We show that the glutamate released from a single or a few release boutons during normal activity acts similarly to glutamate added externally, i.e. it inhibits its own release.
Distribution and Sites of Synthesis of NTT4, an Orphan Member of the Na+/Cl‐‐dependent Neurotransmitter Transporter Family, in the Rat CNS
Authors: Luque J. M. , Jursky F. , Nelson N. , Richards J. G.
Published: April 2006

The distribution and sites of synthesis in rat CNS of NTT4, a novel orphan member of the Na+/Cl‐‐dependent neurotransmitter transporter family, were determined by immunohistochemistry and hybridization histochemistry. Antibodies raised against recombinant fusion proteins, corresponding to residues of NTT4, and 35S‐labelled oligodeoxyribonucleotide probes, were used to delineate the cellular distribution of the transporter at the protein and mRNA levels. High levels of immunoreactivity (mainly in the neuropil) were found in the olfactory bulb, cerebral cortex, striatum, hippocampus, thalamus, substantia nigra, pontine nuclei, cerebellum and spinal cord. The lowest levels were associated with the lateral hypothalamic area and deep mesencephalic nuclei. In situ hybridization signals correlated well with the immunoreactivity, and demonstrated a widespread distribution of NTT4 transcripts exclusively in neurons. NTT4 transcripts appeared widely codistributed with the N‐methyl‐D‐aspartate receptor subunit 1 (1–4b), i.e. spliced variants characterized by a common 5’63 bp insertion. These results indicate that the transporter was associated with neuronal processes in specific glutamate innervated CNS regions. Although the substrate transported by NTT4 remains unknown, our findings suggest a possible role for this carrier protein in glutamate/glycine neurotransmission.
DCG‐IV Selectively Attenuates Rapidly Triggered NMDA‐induced Neurotoxicity in Cortical Neurons
Authors: Buisson Alain , Yu Shan Ping , Choi Dennis W.
Published: April 2006

Molecular cloning has revealed the existence of at least eight subtypes of metabotropic glutamate receptors (mGluRs). We examined the effect of (2S, 1′R, 2′R, 3′R)‐2‐(2, 3‐dicarboxycyclopropyl)glycine (DCG‐IV), a selective agonist of the mGluR 2/3 subtype, on excitotoxicity in mouse cortical cell cultures. Addition of DCG‐IV to the exposure medium partially attenuated the rapidly triggered excitotoxic death induced by a 5 min exposure to 200 μM NMDA. This neuroprotective effect was reversed by coapplication of α‐methyl‐4‐carboxyphenylglycine (MCPG), an antagonist of mGluRs, by pertussis toxin pretreatment and also by preincubation with dibutyryl cAMP, a stable analogue of cAMP. These results suggest that the activation of mGluR 2/3 is neuroprotective in our system. However, DCG‐IV did not attenuate the slowly triggered neuronal death induced by 24 h exposure to low concentrations of NMDA, α‐amino‐1, 3‐cyclopentanedicarboxylic acid (AMPA) or kainate. The failure of DCG‐IV to block slowly triggered NMDA neurotoxicity is likely due to weak NMDA agonist activity, as demonstrated in whole‐cell recording.
Rab3 Proteins and SNAP‐25, Essential Components of the Exocytosis Machinery in Conventional Synapses, are Absent from Ribbon Synapses of the Mouse Retina
Authors: Grabs D. , Bergmann M. , Urban M. , Post A. , Gratzl M.
Published: April 2006

GTP‐binding rab proteins, present in synaptic vesicles and endocrine secretory granules, have been shown to be involved in the control of regulated exocytosis. We found rab3 proteins in immunoblots of diverse areas of the mouse central nervous system (spinal cord, olfactory bulb, hippocampus, cerebellum and neocortex). Immunohistochemical observations at light‐ and electron‐microscopical levels in the hippocampus and other areas revealed rab3 proteins in virtually all synaptic fields and terminals of the areas investigated. In the retina, rab3A immunoreactivity was confined to the inner and outer plexiform layers. Ultrastructural examination revealed that rab3A was present in conventional terminals in the inner plexiform layer and in horizontal cell processes of the outer plexiform layer. In contrast ribbon synapses, which play a key role in transferring information from the photoreceptor cells to the central nervous system, were immunonegative. We also tested whether other proteins of the rab3 family are present in ribbon synapses. However, using an antibody recognizing rab3B and rab3C in addition to rab3A, we found no immunoreactivity in these synapses. Interestingly, we observed also no immunoreactivity for synaptosomal‐associated protein 25 (SNAP‐25) in ribbon synapses, but conventional synapses and horizontal cell processes were heavily stained. Our data show that the known rab3 and SNAP‐25 isoforms, which are components of the secretory apparatus of conventional synapses, are absent from ribbon synapses of the retina. Our observations suggest different mechanisms of transmitter exocytosis in conventional and ribbon terminals.
Potassium Channel mRNA Expression in Prevertebral and Paravertebral Sympathetic Neurons
Authors: Dixon Jane E. , McKinnon David
Published: April 2006

The expression of eighteen different voltage‐activated potassium channel genes in rat sympathetic ganglia was quantitatively analysed using an RNase protection assay. Eleven α‐subunit genes and two β‐subunit genes were expressed in sympathetic ganglia. The relative level of potassium channel mRNA expression was compared between the superior cervical ganglion (SCG) and two prevertebral sympathetic ganglia, the coeliac ganglion (CG) and the superior mesenteric ganglion (SMG). Four mRNAs were differentially expressed: Kv1.2, Kv1.4, Kv2.2 and Kvβ1. Transcripts from all four genes were more abundant in the prevertebral ganglia. From comparisons with previous electrophysiological studies it was concluded that genes encoding the channels underlying the M‐current and D2‐current, which are both prominent in sympathetic neurons, have yet to be identified. It was also concluded that members of the Kv4 family are likely to underlie the low‐threshold A‐current in sympathetic neurons.
Assumptions and Validations of Statistical Tests for Functional Neuroimaging
Authors: Roland P. E. , Gulyas B.
Published: April 2006

We contrast two statistical methods: three‐dimensional cluster analysis and statistical parametric mapping. We show that three‐dimensional cluster analysis is based on a neurobiological theory of the regulation of blood flow and, unlike statistical parametric mapping, carries a minimum of assumptions that are tested. Statistical parametric mapping is a formal approach, which is based on a multitude of assumptions of which the majority have not been validated. We also demonstrate that in practice three‐dimensional cluster analysis has a reasonable balance between sensitivity and the probability of false positives, giving high reproducibility with data on e.g. colour discrimination.
The Reinforcing Properties of Nicotine are Associated with a Specific Patterning of c‐fos Expression in the Rat Brain
Authors: Pagliusi Sonia R. , Tessari Michela , DeVevey Sabine , Chiamulera Christian , Pich Emilio Merlo
Published: April 2006

Rats were trained for nicotine intravenous infusions in a self‐administration paradigm. The effect of nicotine self‐administration on regional brain activity was studied by mapping changes of c‐fos expression. Specific nicotine effects were determined by comparing the patterning of Fos‐like immunoreactivity (Fos‐LI) in nicotine self‐administering rats with that in three different control groups. Controls included rats exposed to the same manipulation as nicotine self‐administering rats who received intravenous saline instead of nicotine. In addition, two groups of untrained sham‐operated rats exposed daily to the same operant boxes were included: one group had the same food restriction used in the operant training, the other was fed ad libitum. Nicotine self‐administration, exposure to saline and food restriction increased Fos‐LI in 43, 33 and three brain regions, respectively, when compared with the control group fed ad libitum. Computer‐assisted image analysis of Fos‐LI profiles performed on 16 relevant limbic and sensory structures showed that in saline‐treated rats a significant (P < 0.01) increase of Fos‐LI profiles was observed in medial prefrontal cortex, lateral septum, core and ventral shell of nucleus accumbens, claustrum, amygdaloid nuclei, paraventricular thalamic nucleus and lateral geniculate nucleus. A significant (P < 0.01) further increase produced by nicotine was found in medial prefrontal cortex and ventral shell of nucleus accumbens. Interestingly, cingulate and piriform cortex, superior colliculus and medial terminal nucleus of the accessory optic tract were specifically activated by nicotine but not saline. These results show that nicotine self‐administration activates sensory structures, as well as limbic structures involved in natural rewarding pathways. The results suggest the involvement of restricted terminal regions of the mesocorticolimbic dopaminergic system in the maintenance of nicotine self‐administration.
Long‐term Induction of Haem Oxygenase‐1 (HSP‐32) in Astrocytes and Microglia Following Transient Focal Brain Ischaemia in the Rat
Authors: Koistinaho Jari , Miettinen Susanna , Keinänen Riitta , Vartiainen Nina , Roivainen Reina , Laitinen Jarmo T.
Published: April 2006

Haem oxygenase‐1 (HO‐1) is a stress protein and a rate‐limiting enzyme in haem degradation, generating ferrous iron, carbon monoxide and bile pigments. HO‐1 has been suggested to be protective against oxidative stress. In the normal rodent brain the enzyme is localized in selected neuron populations, but heat shock, glutathione depletion in vivo and oxidative stress in vitro induce HO‐1 predominantly in glial cells. We studied HO‐1 expression in the brain following transient occlusion of the middle cerebral artery, and found increased mRNA levels in the ischaemic region from 4 h to 7 days after 90 min of ischaemia. The mRNA levels peaked at 12 h, and were localized perifocally. HO‐1‐immunoreactive astrocytes and microglial cells were seen in the perifocal area, in the ipsilateral and occasionally in the contralateral hippocampus. Some perifocal neurons were also HO‐14mmunoreactive. In the infarcted area HO‐1‐positive microglia/macrophages were detected in double‐labelling experiments. A microassay measuring the conversion of [14C]haem to [14C]bilirubin showed a two‐fold increase in haem oxygenase activity in the infarcted core. These observations show a long‐term induction of HO‐1 protein and its activity following ischaemia‐reperfusion brain injury, and indicate increased capacity for haem degradation and the generation of biologically active bile products, carbon monoxide and iron in astrocytes and some microglia/macrophages during focal brain ischaemia.
Distribution of GABAergic Elements Postsynaptic to Ventroposteromedial Thalamic Projections in Layer IV of Rat Barrel Cortex
Authors: Staiger Jochen F. , Zilles Karl , Freund Tamás F.
Published: April 2006

The spatial synaptic pattern formed by boutons, originating in the ventroposteromedial thalamic nucleus, with GABAergic neurons in the rat barrel cortex was mapped. The aim was to shed light on the structural basis by which inhibitory circuits may be activated at the first stage of cortical information processing. The thalamic afferent projection was labelled by anterograde transport of Phaseolus vulgaris leucoagglutinin (PHA‐L), whereas the GABAergic targets in layer IV of the rat barrel cortex were visualized by postembedding GABA immunogold‐labelling or by pre‐embedding parvalbumin immunocytochemistry. In the first set of experiments, we mapped barrels, contained in single ultrathin sections, by means of a computer‐controlled electron microscope stage in their entire layer IV representation. From a total of 1199 asymmetric PHA‐L‐labelled synapses, only 98 were on GABAergic elements, mainly on dendritic shafts. This corresponded to 8.2% of all synapses counted. These synapses on GABAergic targets were essentially homogeneously distributed without a reliable relationship to barrel subdivisions, i.e. hollow versus wall; or layer IVa versus layer IVb. In the second part of the study, we demonstrated that parvalbumin‐containing neurons represent the major GABAergic cell type targetted by thalamic afferents in layer IV of the barrel cortex, since all parvalbumin‐positive cells investigated received multiple synaptic contacts (up to eight synapses per neuron) from the ventroposteromedial thalamic nucleus. These results imply that interneurons responsible for perisomatic inhibition (basket and chandelier cells known to contain parvalbumin) are likely to be strongly excited by thalamic afferents, despite the relatively low proportion of thalamic synapses on GABAergic elements compared to spines of principal cells, and participate in the early stages of cortical sensory information processing.
Neuronal Precursors in the Postnatal Mouse Cerebellum are Fully Committed Cells: Evidence from Heterochronic Transplantations
Authors: Jankovski Aleksandar , Rossi Ferdinando , Sotelo Constantino
Published: April 2006

Neural progenitors are thought to be multipotent cells whose adult phenotype is determined by extrinsic influences acting during and immediately after their last mitosis. To test this hypothesis, postnatal cerebellar precursor cells were placed in the heterochronic cellular environment of the embryonic mouse cerebellar anlage and the resulting phenotypes were determined. To identify the cells arising from postnatal precursors, tissue fragments taken from 3‐ to 8–day‐old cerebellum of several transgenic mouse lines (each expressing the lacZ reporter gene in different sets of neuronal populations) were mixed with fragments taken from the wild‐type cerebellar primordium of 12‐ or 13–day‐old embryos. The fragments were dissociated and grafted into the cerebellum of adult mice. The phenotype acquired by postnatal precursors in the mixed grafts was determined by their morphology and ultrastructural features and by the expression of specific markers. Only two adult phenotypes were generated by these precursors: granule cells and molecular layer interneurons. Most granule cells were well integrated in the trilaminated cortex of the graft, being positioned in their proper layer both during development and after complete maturation. By contrast, basket and stellate cells were always ectopic, remaining outside the molecular layer. These results indicate that at least two distinct progenitor cells are present in the postnatal cerebellar cortex under the experimental conditions of this study. Both progenitors appear to be strictly specified at the time of grafting, and neither their identity nor the expression of their major distinctive features are significantly influenced by local signals emerging from the cellular environment of the embryonic cerebellar anlage.
Induction of Dopaminergic Neurotransmitter Phenotype in Rat Embryonic ‐Cerebrocortex by the Synergistic Action of Neurotrophins and Dopamine
Authors: Zhou Jiawei , Bradford Henry F. , Stern Gerald M.
Published: April 2006

Neurotrophins, including nerve growth factor, brain‐derived neurotrophic factor (BDNF), neurotrophin‐3 (NT‐3) and neurotrophin‐4/51 (NT‐4/5), have been shown to enhance survival and differentiation of a variety of central neuronal populations, such as those with the dopaminergic, cholinergic, GABAergic phenotype during development. In this paper we present evidence that BDNF, NT‐3 and NT‐4/5 acting synergistically with dopamine (DA) can artificially induce the full dopaminergic phenotype in rat fetal cerebral cortex which normally has very few dopaminergic neurons in adulthood. Thus, BDNF/DA, NT‐3/DA, NT‐4/DA elicited a great increase in the number of tyrosine hydroxylase (TH)‐immunoreactive cells, which was up to 57% of total neuronal population in cultures of fetal rat cortical cells. This stimulatory effect was not dependent on glial proliferation, or on addition of serum to the culture. Pharmacological studies showed that dopamine receptors D, and D2 were involved in this effect. The TH’cortical cells possessed other biochemical phenotypic features of dopaminergic neurons. Thus, high‐affinity DA uptake was elevated in cortical cultures treated with neurotrophin/DA. Also DA and 3,4–dihydroxyphenlacetic acid production was detected (5.42 ± 1.24 and 13.72 ? 2.84 pmol/dish respectively, zero in controls). This shows the presence of functionally active TH, aromatic acid decarboxylase and monoamine oxidase. Neurotrophins/DA had no effect on noradrenergic phenotype expression by cortical fetal cells. Taken together, these results support the long‐standing view that development of the central nervous system is determined not only by intrinsic genetic programmes, but also involves environmental influences such as the action of growth factors and extracellular neurotransmitters. In this case we report the effect of specific DA phenotype‐inducing agents.
5‐HT1A Receptor Agonist Flesinoxan Enhances Fos Immunoreactivity in Rat Central Amygdala, Bed Nucleus of the Stria Terminalis and Hypothalamus
Authors: Compaan J. C. , Groenink L. , van der Gugten J. , Maes R. A. A. , Olivier B.
Published: April 2006

5‐Hydroxytryptamine‐1A (5‐HT1A) receptor agonists, including flesinoxan, reduce anxiety and activate the hypothalamus‐pituitary‐adrenal (HPA) axis under basal conditions. In order to investigate the underlying neural mechanisms we investigated immunoreactivity for the immediate early gene protein product Fos (Fos‐ir) in rat brains 1 h after flesinoxan treatment (0.0, 0.3 or 3.0 mg/kg p.0.). Typically, 5‐HT1A receptor‐containing brain areas, such as the dorsal raphe nuclei, hippocampus, septum, diagonal band and the cortical and basomedial amygdala, do not show Fos‐ir. Apparently, binding of flesinoxan at the 5‐HT1A receptor does not directly lead to activation of c‐fos in the cell, probably due to its negative coupling to adenylate cyclase. However, in typically non‐5HTlA receptor‐containing brain areas Fos‐ir is increased due to flesinoxan treatment, as in the paraventricular nucleus of the hypothalamus (PVN), the dorsolateral part of the bed nucleus of the stria terminalis (BNSTd1) and the central amygdala (CeA). Flesinoxan‐treated rats also exhibited higher plasma corticosterone levels than vehicle‐treated animals, which suggests the involvement of corticotropin‐releasing hormone (CRH) or vasopressin in the hypothalamus. After double immunolabelling (Fos/CRH or Fos/vasopressin), every CRH neuron detected in the PVN also contained Fos. Moreover, a significant correlation existed between the number of Fos‐ir neurons in the PVN and the plasma corticosterone level. Hardly any Fos/ vasopressin double labelling was visible in the PVN. Accordingly, flesinoxan exerts its activating effects on the HPA axis via CRH neurons in the PVN. These effects are transsynaptically mediated by other brain areas, such as the CeA and BNSTdl, which also show increased Fos‐ir.
Role of Thyroid Hormone in the Morphological Differentiation and Maturation of Astrocytes: Temporal Correlation with Synthesis and Organization of Actin
Authors: Paul Surojit , Das Sumantra , Poddar Ranjana , Sarkar Pranab Kumar
Published: April 2006

Morphological changes and the molecular mechanisms associated with the maturation of astrocytes were studied under normal and thyroid hormone‐deficient conditions using long‐term (30 days) primary cultures derived from the neonatal rat brain. Immunocytochemical staining of cells with a monoclonal antibody specific to glial fibrillary acidic protein demonstrated for the first time that, similar to their maturation in vivo, astrocytes maintained in normal serum‐containing medium can undergo complete maturation involving two distinct stages of morphological differentiation (from radial glia to flat polygonal cells with epithelioid morphology and then to mature process‐bearing cells with stellate morphology). Deficiency of thyroid hormone delays the first step and totally blocks the second stage of differentiation in the maturation process. Comparative staining of normal and thyroid hormone‐deficient astrocytes with filamentous actin‐specific fluorescein isothiocyanate‐phalloidin and quantitation of the various forms of intracellular actin using an improved DNase I assay demonstrated that maturation of astroglial cells is associated with characteristic alterations in the level of cytoskeletal and non‐cytoskeletal filamentous (F) actin. In particular, the maintenance of the epithelioid form of the hypothyroid astrocytes is associated with a progressive increase in the level of cytoskeletal F‐actin and a concomitant decline in the level of non‐cytoskeletal F‐actin. Quantitation of actin mRNA by Northern blot analysis and studies on the rate of actin synthesis at various stages of differentiation showed that the initial transformation into the epithelioid form is associated with an increase in the rate of synthesis of actin and the expression of its mRNA, while the final transformation into the mature process‐bearing form is correlated with a decline in these parameters. The results indicate that thyroid hormone plays an obligatory role in promoting the differentiation and maturation of astrocytes, and that during this process the hormone regulates the expression of actin and its intracellular organization in a way conducive to morphological differentiation.
The Calretinin‐containing Mossy Cells Survive Excitotoxic Insult in the Gerbil Dentate Gyrus. Comparison of Excitotoxicity‐induced Neuropathological Changes in the Gerbil and Rat
Authors: Kotti T. , Tapiola T. , Riekkinen P. J. , Miettinen R.
Published: April 2006

Our preliminary results showed that mossy fibres do not undergo sprouting after global ischaemia in gerbils, although the pattern of hippocampal cell damage resembled that seen in ischaemic and epileptic rats, where mossy fibre sprouting is known to occur. In order to investigate whether the observed differences in the appearance of mossy fibre sprouting are related to the animal model or species used, this study was undertaken to compare the neuropathological changes induced in gerbils by systemic injection of kainate or by occlusion of carotid arteries with the changes induced in rats by injection of kainate. The pattern of pyramidal cell damage was very similar in each group. Mossy fibre sprouting was present in epileptic rats but not in ischaemic or epileptic gerbils. The number of somatostatin‐immunoreactive neurons was decreased in the hilus of epileptic rats and ischaemic gerbils, but not in epileptic gerbils. The analysis of calretinin immunoreactivity in the dentate gyrus revealed differences between the rat and gerbil. The most striking difference between these species was that the mossy cells contained calretinin in gerbils but not in rats. Cell counting showed that the calretinin‐containing mossy cells had survived both in epileptic and ischaemic gerbils. Therefore, since the mossy cells are known to be highly susceptible to excitotoxic insult in rats and degeneration of these cells is thought to be a key element in the induction of mossy fibre sprouting, we propose that the absence of mossy fibre sprouting in gerbils is related to the survival of the mossy cells.
Expression of Kainate‐selective Glutamate Receptor Subunits in Glial Cells of the Adult Bovine White Matter
Authors: Garcia‐Barcina Jose M. , Matute Carlos
Published: April 2006

It is now well established that glial cells, like neurons, express a large variety of non‐NMDA glutamate receptors. However, knowledge about the subunits forming the receptors and the types of glial cells which express them is incomplete. The aim of this study was to determine the presence and distribution of kainate‐preferring glutamate receptor subunits in glial cells in the white matter of adult cattle. To this end, reverse transcription and polymerase chain reaction (RT‐PCR) analysis of mRNA extracted from the corpus callosum, fornix and optic nerve of brains of adult cattle was carried out using specific primers that allowed selective amplification of the kainate transcripts encoding the GluR5, GluR6, GIuR7, KAI and KA2 subunits. In the three areas studied, transcripts for all five subunits were detected. Restriction digestion of the amplified products confirmed the specificity of the PCR amplifications and showed that the extent of Q/R editing in the GluR6 subunit was lower than that described in the rat whole brain. Immunohistochemical experiments revealed that GluR5‐7 and KA2 subunits appeared in ‐50% of astrocytes and 40% of oligodendrocytes. In astrocytes the receptor subunits were located throughout the cell body and processes, whereas in oligodendrocytes they were mostly restricted to the soma. Double‐labelling studies demonstrated that in the white matter areas studied GluR5‐7 and KA2 subunits are coexpressed in the majority of macroglial cells endowed with kainate receptors. In addition, astrocytes, but not oligodendrocytes, expressing kainate‐selective receptors were found to possess the GluR1 subunit, an γ‐amino‐3‐hydroxyd‐methyl‐4‐isoxazole propionic acid (AMPA) receptor subunit which has previously been found in this macroglial cell type. Taken together, these results provide evidence for the presence and molecular diversity of kainate‐selective receptor subunits in glial cells of the adult white matter and for their co‐expression with AMPA receptor subunits. The widespread distribution of kainate receptors in glial cells of the white matter suggests that these receptors may be important in axon‐to‐glia and/or glia‐to‐glia signalling.
The G‐protein γ‐subunit Gy8 is Expressed in the Developing Axons of Olfactory and Vomeronasal Neurons
Authors: Tirindelli Roberto , Ryba Nicholas J. P.
Published: April 2006

The tissue localization of the G‐protein y‐subunit, Gy8, that is specifically expressed in the olfactory and vomeronasal neurons, was studied in rats at different ages: embryonic day 16, postnatal days 1,7, 14 and 35, and adult. G8 appears to be a specific marker of the immature olfactory and vomeronasal neurons. Its distribution differs from that of Golfα, a G‐protein α‐subunit which is predominantly expressed in mature olfactory neurons. G8 immunoreactivity indicates that an undifferentiated organization of the olfactory epithelium persists up to 3 weeks of age, though neonates possess a functional sense of smell. Gy8 accumulates at the highest levels in the axons of the developing olfactory neurons 2 weeks after birth (postnatal day 14). Moreover, up to postnatal day 14, Gy8–positive neurons are present in the region of the olfactory and vomeronasal epithelium, where they are not observed in later life. In the olfactory epithelium and in the bulb, Gy8 expression becomes weaker and patchy with increasing age, suggesting that the process of continuous regeneration of olfactory neurons occurs in discrete areas. G8–enhanced expression following axotomy indicates that this system is potentially active throughout life. Conversely, in the vomeronasal epithelium Gy8 expression persists virtually unmodified in the adult. This indicates that the degree of differentiation may differ between olfactory and vomeronasal neurons.
Timing and Regulation of trkB and BDNF mRNA Expression in Placode‐derived Sensory Neurons and their Targets
Authors: Robinson Michelle , Adu Jimi , Davies Alun M.
Published: April 2006

The sensory neurons of the vestibular and nodose ganglia of the chicken embryo have nearby and distant targets, respectively. in vitro studies have shown that these neurons survive independently of neurotrophins when their axons are growing to their targets and become dependent on brain‐derived neurotrophic factor (BDNF) for survival when their axons reach the vicinity of their targets. Although the timing of BDNF dependence is principally controlled by an intrinsic timing mechanism in the neurons, the onset of dependence can be accelerated by BDNF exposure toward the end of the phase of neurotrophin independence. We have used quantitative reverse transcription/polymerase chain reaction to study the expression of transcripts coding for BDNF and the catalytic isoform of its receptor tyrosine kinase, TrkB, in these neurons and their targets at different stages of development. We show that the peripheral and central target tissues of these neurons express BDNF mRNA prior to the arrival of sensory axons. Vestibular neurons express trk6 mRNA before nodose neurons, which accords with the earlier response of vestibular neurons to BDNF. In culture, early nodose neurons start expressing trkB mRNA after 36 h incubation, which is 36 h before these neurons become dependent on BDNF for survival. Although BDNF does not affect the timing and level of trkB mRNA expression during the first 48 h in vitm, it increases the level of trk8 mRNA after this time. The timing of BDNF‐induced elevation of trkB mRNA correlates with the period during which BDNF exposure accelerates the onset of BDNF dependence in nodose neurons. These results suggest that the timing of BDNF dependence in developing sensory neurons is due in part to expression of catalytic TrkB and demonstrate that a BDNF autocrine loop is not required for the survival of sensory neurons during the earliest stages of their development.
Immortalized Hypothalamic Neurons Express Metabotropic Glutamate Receptors Positively Coupled to Cyclic AMP Formation
Authors: Sortino M. A. , Aleppo G. , Copani A. , Casabona G. , Nicoletti F. , Ventra C. , Kuhn R. , Knöpfel T. , Malitschek B. , Canonico P. L.
Published: April 2006

We have characterized the expression pattern and pharmacological profile of activation of metabotropic glutamate receptors (mGluRs) in immortalized, gonadotropin releasing hormone (GnRH)‐secreting GT1‐7 cells, which represent a homogeneous cellular population of hypothalamic origin. These cells are known to respond to the mGluR agonist (1S, 3R)‐cyclopentanedicarboxylic acid (1S, 3R‐ACPD) with increased GnRH release. To establish which specific mGluR subtypes are expressed by GT1‐7 cells, we used polyclonal antibodies raised against non‐conserved regions of the carboxy‐terminal domains of individual subtypes. The selectivity of these antibodies was tested in HEK 293 cells transiently transfected with each mGluR subtype. GTl‐7 cells stained positively for the subtypes mGluRla, ‐1b and ‐5 (belonging to group I mGluRs), mGluR2/3 (group 11) and mGluR7 (group 111). Agonists of group I mGluRs, including 1S, 3R‐ACPD, activated phosphoinositide hydrolysis in GT1‐7 cells. This effect, however, was manifested only when cell density was low, and it disappeared when cells reached confluence. Stimulation of phosphoinositide hydrolysis could not therefore have been related to hormone secretion because 1S, 3RACPD effectively released GnRH in confluent cultures. We then focused on group II and Ill mGluRs, which in transfected cells are negatively linked to adenylate cyclase activity. Unexpectedly, however, agonists which preferentially activate group II and Ill mGluRs increased both basal and forskolin‐stimulated cAMP accumulation in GT1‐7 cells. Stimulation of GAMP accumulation by mGluR agonists was not prevented by enzymatic depletion of endogenous adenosine, but was obliterated when cells were incubated with agonists of receptors positively coupled to adenylate cyclase, such as P‐adrenergic and prostaglandin E2 receptors. These results suggest that GT1‐7 cells express a novel mGluR subtype positively coupled to adenylate cyclase, which shares the same transduction pathway of other classical receptors coupled with a G,‐type of GTP‐binding protein.
The Course and Termination of Corticothalamic Fibres Arising in the Visual Cortex of the Rat
Authors: Lozsádi D. A. , Gonzalez‐Soriano J. , Guillery R. W.
Published: April 2006

Corticothalamic axons have been studied in adult Lister hooded rats with single or dual injections of tracers into the visual cortex. Labelled axons leave medial and lateral injection sites in separate or partially overlapping bundles along parallel trajectories in the subcortical white matter. In the internal capsule they converge and both bundles enter roughly the same sector of the thalamic reticular nucleus (TRN). Their reticular terminal fields, however, differ. Axons from a medial injection site innervate more lateral parts of the TRN than do the axons from lateral injection sites. The most medial third of the TRN is not innervated from area 17 but receives a topographically arranged input from peristriate cortex (Crabtree and Killackey, 1989, Eur J. Neurosci., 1, 94‐109; Coleman and Mitrofanis, 1996, EWK J. Neurosci., 8, 388‐404). The two groups of axons then separate in the dorsal thalamus, axons from medial parts of visual cortex turning caudally into lateral regions of the lateral geniculate nucleus, whereas fibres from more lateral cortex continue into medial parts of the nucleus. Connolly and van Essen (1984, J. Comp. Neurol., 226, 544‐564) and Nelson and LeVay (1985, J. Comp. Neurol., 240, 322‐330) have shown that in the geniculocortical pathway the two groups of fibres cross over in the subcortical white matter, probably in the region of the subplate. We show that the corticothalamic pathway also has a crossing, but it occurs in, or close to, the diencephalon itself, in the region of the perireticular nucleus. This result suggests that each of these pathways, the geniculocortical and the corticogeniculate, may undergo reorganization within distinct cerebral zones, one diencephalic for the corticothalamic axons and the other telencephalic for the thalamocortical axons.