bluuebunny/crossref_metadata_2025 huggingface dataset:
Biochemical and enzymological aspects of the symbiosis between the deep‐sea tubeworm Riftia pachyptila and its bacterial endosymbiont
Authors: Minic Zoran , Hervé Guy
Published: July 2004

Riftia pachyptila (Vestimentifera) is a giant tubeworm living around the volcanic deep‐sea vents of the East Pacific Rise. This animal is devoid of a digestive tract and lives in an intimate symbiosis with a sulfur‐oxidizing chemoautotrophic bacterium. This bacterial endosymbiont is localized in the cells of a richly vascularized organ of the worm: the trophosome. These organisms are adapted to their extreme environment and take advantage of the particular composition of the mixed volcanic and sea waters to extract and assimilate inorganic metabolites, especially carbon, nitrogen, oxygen and sulfur. The high molecular mass hemoglobin of the worm is the transporter for both oxygen and sulfide. This last compound is delivered to the bacterium which possesses the sulfur oxidizing respiratory system, which produces the metabolic energy for the two partners. CO2 is also delivered to the bacterium where it enters the Calvin–Benson cycle. Some of the resulting small carbonated organic molecules are thus provided to the worm for its own metabolism. As far as nitrogen assimilation is concerned, NH3 can be used by the two partners but nitrate can be used only by the bacterium. This very intimate symbiosis applies also to the organization of metabolic pathways such as those of pyrimidine nucleotides and arginine. In particular, the worm lacks the first three enzymes of the de novo pyrimidine biosynthetic pathways as well as some enzymes involved in the biosynthesis of polyamines. The bacterium lacks the enzymes of the pyrimidine salvage pathway. This symbiotic organization constitutes a very interesting system to study the molecular and metabolic basis of biological adaptation.
Presence of membrane ecdysone receptor in the anterior silk gland of the silkworm Bombyx mori
Authors: Elmogy Mohamed , Iwami Masafumi , Sakurai Sho
Published: July 2004

Nongenomic action of an insect steroid hormone, 20‐hydroxyecdysone (20E), has been implicated in several 20E‐dependent events including the programmed cell death of Bombyx anterior silk glands (ASGs), but no information is available for the mode of the action. We provide evidence for a putative membrane receptor located in the plasma membrane of the ASGs. Membrane fractions prepared from the ASGs exhibit high binding activity to [3H]ponasterone A (PonA). The membrane fractions did not contain conventional ecdysone receptor as revealed by Western blot analysis using antibody raised against Bombyx ecdysone receptor A (EcR‐A). The binding activity was not solubilized with 1 m NaCl or 0.05% (w/v) MEGA‐8, indicating that the binding sites were localized in the membrane. Differential solubilization and temperature‐induced phase separation in Triton X‐114 showed that the binding sites might be integrated membrane proteins. These results indicated that the binding sites are located in plasma membrane proteins, which we putatively referred to as membrane ecdysone receptor (mEcR). The mEcR exhibited saturable binding for [3H]PonA (Kd = 17.3 nm, Bmax = 0.82 pmol·mg−1 protein). Association and dissociation kinetics revealed that [3H]PonA associated with and dissociated from mEcR within minutes. The combined results support the existence of a plasmalemmal ecdysteroid receptor, which may act in concert with the conventional EcR in various 20E‐dependent developmental events.
α‐Synuclein–synaptosomal membrane interactions
Authors: Jo Euijung , Darabie Audrey A. , Han Kyung , Tandon Anurag , Fraser Paul E. , McLaurin JoAnne
Published: July 2004

α‐Synuclein exists in two different compartments in vivo– correspondingly existing as two different forms: a membrane‐bound form that is predominantly α‐helical and a cytosolic form that is randomly structured. It has been suggested that these environmental and structural differences may play a role in aggregation propensity and development of pathological lesions observed in Parkinson's disease (PD). Such effects may be accentuated by mutations observed in familial PD kindreds. In order to test this hypothesis, wild‐type and A53T mutant α‐synuclein interactions with rat brain synaptosomal membranes were examined. Previous data has demonstrated that the A30P mutant has defective lipid binding and therefore was not examined in this study. Electron microscopy demonstrated that wild‐type α‐synuclein fibrillogenesis is accelerated in the presence of synaptosomal membranes whereas the A53T α‐synuclein fibrillogenesis is inhibited under the same conditions. These results suggested that subtle sequence changes in α‐synuclein could significantly alter interaction with membrane bilayers. Fluorescence and absorption spectroscopy using environment sensitive probes demonstrated variations in the inherent lipid properties in the presence and absence of α‐synuclein. Addition of wild‐type α‐synuclein to synaptosomes did not significantly alter the membrane fluidity at either the fatty acyl chains or headgroup space, suggesting that synaptosomes have a high capacity for α‐synuclein binding. In contrast, synaptosomal membrane fluidity was decreased by A53T α‐synuclein binding with concomitant packing of the lipid headgroups. These results suggest that alterations in α‐synuclein–lipid interactions may contribute to physiological changes detected in early onset PD.
Interaction of selenium compounds with zinc finger proteins involved in DNA repair
Authors: Blessing Holger , Kraus Silke , Heindl Philipp , Bal Wojciech , Hartwig Andrea
Published: July 2004

As an essential element, selenium is present in enzymes from several families, including glutathione peroxidases, and is thought to exert anticarcinogenic properties. A remarkable feature of selenium consists of its ability to oxidize thiols under reducing conditions. Thus, one mode of action recently suggested is the oxidation of thiol groups of metallothionein, thereby providing zinc for essential reactions. However, tetrahedral zinc ion complexation to four thiolates, similar to that found in metallothionein, is present in one of the major classes of transcription factors and other so‐called zinc finger proteins. Within this study we investigated the effect of selenium compounds on the activity of the formamidopyrimidine‐DNA glycosylase (Fpg), a zinc finger protein involved in base excision repair, and on the DNA‐binding capacity and integrity of xeroderma pigmentosum group A protein (XPA), a zinc finger protein essential for nucleotide excision repair. The reducible selenium compounds phenylseleninic acid, phenylselenyl chloride, selenocystine, ebselen, and 2‐nitrophenylselenocyanate caused a concentration‐dependent decrease of Fpg activity, while no inhibition was detected with fully reduced selenomethionine, methylselenocysteine or some sulfur‐containing analogs. Furthermore, reducible selenium compounds interfered with XPA–DNA binding and released zinc from the zinc finger motif, XPAzf. Zinc release was even evident at high glutathione/oxidised glutathine ratios prevailing under cellular conditions. Finally, comparative studies with metallothionein and XPAzf revealed similar or even accelerated zinc release from XPAzf. Altogether, the results indicate that zinc finger motifs are highly reactive towards oxidizing selenium compounds. This could affect gene expression, DNA repair and, thus, genomic stability.
Structural basis of charge transfer complex formation by riboflavin bound to 6,7‐dimethyl‐8‐ribityllumazine synthase
Authors: Koch Michael , Breithaupt Constanze , Gerhardt Haase Stefan, Ilka , Weber Stefan , Cushman Mark , Huber Robert , Bacher Adelbert , Fischer Markus
Published: July 2004

The amino acid residue tryptophan 27 of 6,7‐dimethyl‐8‐ribityllumazine synthase of the yeast Schizosaccharomyces pombe was replaced by tyrosine. The structures of the W27Y mutant protein in complex with riboflavin, the substrate analogue 5‐nitroso‐6‐ribitylamino‐2,4(1H,3H)‐pyrimidinedione, and the product analogue 6‐carboxyethyl‐7‐oxo‐8‐ribityllumazine, were determined by X‐ray crystallography at resolutions of 2.7–2.8 Å. Whereas the indole system of W27 forms a coplanar π‐complex with riboflavin, the corresponding phenyl ring in the W27Y mutant establishes only peripheral contact with the heterocyclic ring system of the bound riboflavin. These findings provide an explanation for the absence of the long wavelength shift in optical absorption spectra of riboflavin bound to the mutant enzyme. The structures of the mutants are important tools for the interpretation of the unusual physical properties of riboflavin in complex with lumazine synthase.
On the mechanism of action of the antifungal agent propionate
Authors: Brock Matthias , Buckel Wolfgang
Published: July 2004

Propionate is used to protect bread and animal feed from moulds. The mode of action of this short‐chain fatty acid was studied using Aspergillus nidulans as a model organism. The filamentous fungus is able to grow slowly on propionate, which is oxidized to acetyl‐CoA via propionyl‐CoA, methylcitrate and pyruvate. Propionate inhibits growth of A. nidulans on glucose but not on acetate; the latter was shown to inhibit propionate oxidation. When grown on glucose a methylcitrate synthase deletion mutant is much more sensitive towards the presence of propionate in the medium as compared to the wild‐type and accumulates 10‐fold higher levels of propionyl‐CoA, which inhibits CoA‐dependent enzymes such as pyruvate dehydrogenase, succinyl‐CoA synthetase and ATP citrate lyase. The most important inhibition is that of pyruvate dehydrogenase, as this affects glucose and propionate metabolism directly. In contrast, the blocked succinyl‐CoA synthetase can be circumvented by a succinyl‐CoA:acetate/propionate CoA‐transferase, whereas ATP citrate lyase is required only for biosynthetic purposes. In addition, data are presented that correlate inhibition of fungal polyketide synthesis by propionyl‐CoA with the accumulation of this CoA‐derivative. A possible toxicity of propionyl‐CoA for humans in diseases such as propionic acidaemia and methylmalonic aciduria is also discussed.
A novel coupled enzyme assay reveals an enzyme responsible for the deamination of a chemically unstable intermediate in the metabolic pathway of 4‐amino‐3‐hydroxybenzoic acid in Bordetella sp. strain 10d
Authors: Orii Chika , Takenaka Shinji , Murakami Shuichiro , Aoki Kenji
Published: July 2004

2‐Amino‐5‐carboxymuconic 6‐semialdehyde is an unstable intermediate in the meta‐cleavage pathway of 4‐amino‐3‐hydroxybenzoic acid in Bordetella sp. strain 10d. In vitro, this compound is nonenzymatically converted to 2,5‐pyridinedicarboxylic acid. Crude extracts of strain 10d grown on 4‐amino‐3‐hydroxybenzoic acid converted 2‐amino‐5‐carboxymuconic 6‐semialdehyde formed from 4‐amino‐3‐hydroxybenzoic acid by the first enzyme in the pathway, 4‐amino‐3‐hydroxybenzoate 2,3‐dioxygenase, to a yellow compound (εmax = 375 nm). The enzyme in the crude extract carrying out the next step was purified to homogeneity. The yellow compound formed from 4‐amino‐3‐hydroxybenzoic acid by this purified enzyme and purified 4‐amino‐3‐hydroxybenzoate 2,3‐dioxygenase in a coupled assay was identified as 2‐hydroxymuconic 6‐semialdehyde by GC‐MS analysis. A mechanism for the formation of 2‐hydroxymuconic 6‐semialdehyde via enzymatic deamination and nonenzymatic decarboxylation is proposed based on results of spectrophotometric analyses. The purified enzyme, designated 2‐amino‐5‐carboxymuconic 6‐semialdehyde deaminase, is a new type of deaminase that differs from the 2‐aminomuconate deaminases reported previously in that it primarily and specifically attacks 2‐amino‐5‐carboxymuconic 6‐semialdehyde. The deamination step in the proposed pathway differs from that in the pathways for 2‐aminophenol and its derivatives.
Calcium‐independent cytoskeleton disassembly induced by BAPTA
Authors: Saoudi Yasmina , Rousseau Bernard , Doussière Jacques , Charrasse Sophie , Gauthier‐Rouvière Cécile , Morin Nathalie , Sautet‐Laugier Christelle , Denarier Eric , Scaïfe Robin , Mioskowski Charles , Job Didier
Published: July 2004

In living organisms, Ca2+ signalling is central to cell physiology. The Ca2+ chelator 1,2‐bis(2‐aminophenoxy)ethane‐N,N,N′,N′‐tetraacetic acid (BAPTA) has been widely used as a probe to test the role of calcium in a large variety of cell functions. Here we show that in most cell types BAPTA has a potent actin and microtubule depolymerizing activity and that this activity is completely independent of Ca2+ chelation. Thus, the depolymerizing effect of BAPTA is shared by a derivative (D‐BAPTA) showing a dramatically reduced calcium chelating activity. Because the extraordinary depolymerizing activity of BAPTA could be due to a general depletion of cell fuel molecules such as ATP, we tested the effects of BAPTA on cellular ATP levels and on mitochondrial function. We find that BAPTA depletes ATP pools and affects mitochondrial respiration in vitro as well as mitochondrial shape and distribution in cells. However, these effects are unrelated to the Ca2+ chelating properties of BAPTA and do not account for the depolymerizing effect of BAPTA on the cell cytoskeleton. We propose that D‐BAPTA should be systematically introduced in calcium signalling experiments, as controls for the known and unknown calcium independent effects of BAPTA. Additionally, the concomitant depolymerizing effect of BAPTA on both tubulin and actin assemblies is intriguing and may lead to the identification of a new control mechanism for cytoskeleton assembly.
The role of the ESSS protein in the assembly of a functional and stable mammalian mitochondrial complex I (NADH‐ubiquinone oxidoreductase)
Authors: Potluri Prasanth , Yadava Nagendra , Scheffler Immo E.
Published: July 2004

The ESSS protein is a recently identified subunit of mammalian mitochondrial complex I. It is a relatively small integral membrane protein (122 amino acids) found in the β‐subcomplex. Genomic sequence database searches reveal its localization to the X‐chromosome in humans and mouse. The ESSS cDNA from Chinese hamster cells was cloned and shown to complement one complementation group of our previously described mutants with a proposed X‐linkage. Sequence analyses of the ESSS cDNA in these mutants revealed chain termination mutations. In two of these mutants the protein is truncated at the C‐terminus of the targeting sequence; the mutants are null mutants for the ESSS subunit. There is no detectable complex I assembly and activity in the absence of the ESSS subunit as revealed by blue native polyacrylamide gel electrophoresis (BN/PAGE) analysis and polarography. Complex I activity can be restored with ESSS subunits tagged with either hemagglutinin (HA) or hexahistidine (His6) epitopes at the C‐terminus. Although, the accumulation of ESSS‐HA is not dependent upon the presence of mtDNA‐encoded subunits (ND1‐6,4 L), it is incorporated into complex I only in presence of compatible complex I subunits from the same species.
Thermodynamic analysis of porphyrin binding to Momordica charantia (bitter gourd) lectin
Authors: Sultan Nabil A. M. , Maiya Bhaskar G. , Swamy Musti J.
Published: July 2004

Owing to the use of porphyrins in photodynamic therapy for the treatment of malignant tumors, and the preferential interaction of lectins with tumor cells, studies on lectin–porphyrin interaction are of significant interest. In this study, the interaction of several free‐base and metalloporphyrins with Momordica charantia (bitter gourd) lectin (MCL) was investigated by absorption spectroscopy. Difference absorp‐tion spectra revealed that significant changes occur in the Soret band region of the porphyrins on binding to MCL. These changes were monitored to obtain association constants (Ka) and stoichiometry of binding. The tetrameric MCL binds four porphyrin molecules, and the stoichiometry was unaffected by the presence of the specific sugar, lactose. In addition, the agglutination activity of MCL was unaffected by the presence of the porphyrins used in this study, clearly indicating that porphyrin and carbohydrate ligands bind at different sites. Both cationic and anionic porphyrins bind to the lectin with comparable affinity (Ka =103−105 m−1). The thermodynamic parameters associated with the interaction of several porphyrins, obtained from the temperature dependence of the Ka values, were found to be in the range: ΔH° = −98.1 to −54.4 kJ·mol−1 and ΔS° =−243.9 to −90.8 J·mol−1·K−1. These results indicate that porphyrin binding to MCL is governed by enthalpic forces and that the contribution from binding entropy is negative. Enthalpy–entropy compensation was observed in the interaction of different porphyrins with MCL, underscoring the role of water structure in the overall binding process. Analysis of CD spectra of MCL indicates that this protein contains about 13%α‐helix, 36%β‐sheet, 21%β‐turn, and the rest unordered structures. Binding of porphyrins does not significantly alter the secondary and tertiary structures of MCL.
Role of Leptin in Farm Animals: a Review
Authors: Máčajová M. , Lamošová D. , Zeman M.
Published: July 2004

The discovery of hormone leptin has led to better understanding of the energy balance control. In addition to its effects on food intake and energy expenditure, leptin has now been implicated as a mediator of diverse physiological functions. Recently, leptin has been cloned in several domestic species. The sequence similarity suggests a common function or mechanism of this peptide hormone across species. Leptin receptors are expressed in most of tissues, which is consistent with the multiplicity of leptin functions. The main goal of this review was to summarize knowledge about effect of leptin on physiology of farm animals. Experiments point to a stimulatory action of leptin on growth hormone (GH) secretion, normal growth and development of the brain. Surprisingly, leptin is synthesized at a high rate in placenta and may function as a growth factor for fetus, signalling the nutritional status from the mother to her offspring. Maturation of reproductive system can be stimulated by leptin administration. Morphological and hormonal changes, consistent with a major role of leptin in the reproductive system, have also been described, including the stimulation of the release of luteinizing hormone (LH), follicle‐stimulating hormone (FSH) and prolactin. Leptin has a substantial effect on food intake and feeding behaviour in animals. Administration of leptin reduces food intake. Its level decrease within hours after initiation of fasting. Leptin also serves as a mediator of the adaption to fasting, and this role may be the primary function for which was the molecule evolved.
A Comparison of Biopsy and Post‐mortem Findings in the Lungs of Healthy Cows
Authors: Sydler T. , Braun U. , Estermann U. , Pospischil A.
Published: July 2004

Lung biopsy in cattle for the diagnosis of lung diseases is rarely used or described. The clinical effects and the gross findings in lung and pleura after ultrasonic‐guided percutaneous lung biopsy in the upright animal of healthy cattle were reported previously (Braun et al., 1999). This report describes the yield of attained lung tissue and the local tissue reaction in 60 healthy adult cows slaughtered 24 h (50 animals) and 10 days (10 animals) after invasion. The yield of lung tissue in the biopsies was high (85.5%) in 90 histologically examined biopsy specimens and judged ‘good to excellent’. The local tissue reaction and the extent of haemorrhage in the lung parenchyma, both 24 h and 10 days after the biopsy, were minimal. In the latter group, the scar tissue had approximately the same dimensions as the tissue cylinders punched 10 days before. There were no local adhesions between the pleural surfaces. Previously published data concerning clinical reactions and complications, macroscopically examined local tissue bleeding and the histologically evaluated minimal local inflammatory reaction, following lung biopsy of cattle, indicate that this procedure is safe and satisfactory for sampling of accurate lung material. The results of this study concur and indicate that local complications in the animal were minimal to absent.
Relationships between Lactational Incidence of Displaced Abomasum and Milk Production Traits in German Holstein Cows
Authors: Hamann H. , Wolf V. , Scholz H. , Distl O.
Published: July 2004

The objective of this study was to analyse the heritabilitiy of the lactational incidence of displaced abomasum (DA) and the relationships of DA with milk production traits in German Holstein cows. Data were recorded between February 1999 and January 2000 in cooperation with five veterinary practitioners. Their veterinary practices were located in the northern part of Lower Saxony. The investigation included 160 dairy farms under the official milk‐recording scheme with 9315 cows. The lactational incidence of the left abomasal displacement amounted to 1.21%, and of the right abomasal displacement to 0.41%, respectively. The linear heritability estimates for the lactational incidences of left and right DA were h2 =0.05 ± 0.012 and h2 = 0.004 ± 0.005, respectively. Using the Dempster–Lerner‐transformation the corresponding heritabilities were h2 = 0.53 and 0.09, respectively. Milk losses for the lactation when DA was diagnosed were significant and reached 1016 kg milk, 41 kg fat, 36 kg protein and 0.07% protein. Fat content significantly increased by 0.18%. The analysis could not show significant differences between cows diagnosed with DA and cows not diagnosed with DA in the 305‐day milk production traits of the lactation preceding the diagnosis of DA. There was also no indication for an unequal distribution of breeding values for milk performance traits between cows with and without DA. The additive genetic correlations between 305‐day milk performance and left DA were low. The results indicated that cows with a high milk production and superior breeding values for milk performance were not exposed to an increased risk for DA.
Identification of a novel brain‐specific and reelin‐regulated gene that encodes a protein colocalized with synapsin
Authors: Kuvbachieva Anelia , Bestel Aude‐Marie , Tissir Fadel , Maloum Ismahane , Guimiot Fabien , Ramoz Nicolas , Bourgeois Francine , Moalic Jean‐Marie , Goffinet André M. , Simonneau Michel
Published: July 2004

We carried out a screening of genes that are differentially expressed in normal mice and reeler mutants and are characterized by abnormal neuronal migration and neurite deployment due to defective Reelin signalling. A novel gene, provisionally named C61, was overexpressed in Reelin‐deficient embryonic mouse brain RNA. C61 encodes a 3.7 kb mRNA that is brain specific and developmentally regulated, with predominant expression in differentiating neurons. The predicted protein is 664 amino acids long, and contains LAG1 and Ezrin/Radixin/Moesin‐Myosin‐Filament motifs, suggesting that it may function as an intracellular adaptor. From E14.5 to birth, C61 was highly expressed in all neuronal differentiation fields, with the highest signal in the telencephalic cortical plate and mitral cells in the olfactory bulb. When expressed as a GFP fusion protein in transfected non‐neuronal cells and primary neurons, this protein localizes, respectively, to the nuclear membrane or axonal outgrowths, indicating a function in axonal traffic or signalling.
Mechanism of GABAB receptor‐mediated inhibition of spontaneous GABA release onto cerebellar Purkinje cells
Authors: Harvey Victoria L. , Stephens Gary J.
Published: July 2004

γ‐Aminobutyric acid (GABA)B receptor‐mediated modulation of spontaneous GABA release onto Purkinje cells was investigated in cerebellar slices from 3‐ to 5‐week‐old mice. The GABAB receptor agonists baclofen and CGP 44533 each reduced the frequency of miniature inhibitory postsynaptic currents (mIPSCs), with no significant effect on mIPSC amplitude; together, consistent with a presynaptic site of action. The GABAB receptor antagonist CGP 55845 blocked baclofen‐induced inhibition. The sulphydryl alkylating agent N‐ethylmaleimide occluded baclofen effects, implicating Gi/o subunits in mediating a GABAB G protein‐coupled receptor pathway. Baclofen‐induced inhibition persisted in the presence of Ba2+, a blocker of K+ channels, and Cd2+, a blocker of Ca2+ channel‐mediated GABA release. Application of nominally Ca2+‐free extracellular solutions reduced mIPSC frequency and amplitude; however, baclofen produced a significant inhibition in mIPSC frequency, further suggesting that this pathway was independent of Ca2+ influx. Spontaneous GABA release was increased by the adenylate cyclase activator, forskolin, and the phorbol ester, phorbol 12,13‐dibutyrate. However, baclofen‐induced inhibition was not significantly changed in either condition. Baclofen action was also not affected by the adenylate cyclase inhibitor SQ 22536 or the protein kinase C inhibitor chelerythrine chloride. Baclofen still reduced mIPSC frequency in the presence of the polyvalent cation ruthenium red, which acts as a secretagogue here; however, baclofen‐induced inhibition was reduced significantly. Furthermore, baclofen produced no clear inhibition during high‐frequency mIPSCs bursts induced by the potent secretagogue α‐Latrotoxin. Together, these results suggest that GABAB inhibition occurs downstream of Ca2+ influx and may be mediated, in part, by an inhibition of the vesicular release mechanism.
Evidence for differential modulation of conditioned aversion and fear‐conditioned analgesia by CB1 receptors
Authors: Finn D. P. , Beckett S. R. G. , Richardson D. , Kendall D. A. , Marsden C. A. , Chapman V.
Published: July 2004

Fear‐conditioned analgesia is an important survival response mediated by substrates controlling nociception and aversion. Cannabinoid1 (CB1) receptors play an important role in nociception and aversion. However, their role in fear‐conditioned analgesia has not been investigated. This study investigated the effects of systemic administration of the CB1 receptor antagonist, SR141716A (1 mg/kg, ip), on fear‐conditioned analgesia and conditioned aversion in rats. Twenty‐four hours after receiving footshock, rats exhibited reduced formalin‐evoked nociceptive behaviour, increased freezing and increased defecation when tested in the footshock apparatus, compared with non‐footshocked formalin‐injected rats. SR141716A attenuated fear‐conditioned analgesia, freezing and defecation. Importantly, SR141716A had no effect on formalin‐evoked nociceptive behaviour over an equivalent time period in rats not receiving footshock. SR141716A had no effect on contextually induced freezing during the first half of the test trial in rats receiving intra‐plantar injection of saline. Administration of SR1417176A did, however, attenuate short‐term extinction of contextually induced freezing and ultrasound emission in rats receiving intra‐plantar saline, compared with vehicle‐treated saline controls. These data suggest an important role for the CB1 receptor in mediating fear‐conditioned analgesia and provide evidence for differential modulation of conditioned aversive behaviour by CB1 receptors during tonic, persistent pain.
Subcortical visual system of the African mole‐ratCryptomys anselli: to see or not to see?
Authors: Němec Pavel , Burda Hynek , Peichl Leo
Published: July 2004

We studied the retinal projections, the distribution of cytochrome oxidase activity and the cyto‐ and myeloarchitecture of the subcortical visual system in the subterranean Ansell's mole‐ratCryptomys anselli. The optic nerve contained 1500 myelinated and a similar number of unmyelinated fibres. The retina projected to all the visual structures described in surface‐dwelling sighted rodents. The suprachiasmatic nucleus was large and received bilateral retinal input. All other visual nuclei were reduced in size, were cytoarchitecturally poorly developed and received almost exclusively contralateral retinal projections. The dorsal and ventral lateral geniculate nuclei were moderately reduced and heavily innervated. The intergeniculate leaflet could be identified between these two nuclei. Pretectal nuclei were also relatively well‐developed. The nucleus of the optic tract, the olivary pretectal nucleus, and the anterior and posterior pretectal nuclei were innervated by the retina. By contrast, the superficial, retinorecipient layers of the superior colliculus showed extreme reduction. The strata zonale, griseum superficiale and opticum were collapsed to a single layer 40 µm thick. The accessory optic system was vestigial. These findings indicate that the functional subsystems involved in photoperiod perception, form and brightness discrimination, and movement analysis are anatomically rather well developed, whereas those involved in coordination of visuomotor reflexes are severely reduced. Thus, the visual system ofC. anselliis much better developed than that of the blind mole‐ratSpalax ehrenbergi.We suggest thatCryptomys ansellihas retained basic visual capabilities.
Neurotrophins induce short‐term and long‐term changes of cortical neurotrophin expression
Authors: Patz Silke , Wahle Petra
Published: July 2004

Neuronal activity, hormones, transmitters, physical exercise and enrichment influence cortical neurotrophin expression. Neurotrophins then elicit structural and physiological changes, and regulate gene expression. This prompted the hypothesis that neurotrophins themselves are involved in regulating neurotrophin expression. Here we investigated the mRNA expression level of brain‐derived neurotrophic factor (BDNF), neurotrophin‐4 (NT‐4), NT‐3 and nerve growth factor (NGF) as well as the tyrosine receptor kinases TrkB and TrkC receptor in response to BDNF, NT‐4, NT‐3 and NGF pulses in organotypic cortex cultures. Single neurotrophin pulses evoked a dramatic up‐ or down‐regulation of some, but not all four, neurotrophin mRNAs, even within 3–24 h, indicating an immediate impact on neurotrophin transcription. Most strikingly, neurotrophin pulses during the first 10 days in vitro (DIV) potentiated the expression of some neurotrophin mRNAs at 20 DIV, suggesting that early trophic factor experience influences the expression levels seen later in development. The NT‐3 mRNA expression, for example, was consistently promoted by NGF and BDNF, suggesting that these two factors help to maintain the low level of NT‐3 found in adult cortex. Rapid bidirectional changes characterized the NT‐4 mRNA expression. A single pulse of NT‐4 transiently increased NT‐4 mRNA, whereas a BDNF pulse transiently reduced NT‐4 mRNA. Surprisingly, NGF strongly potentiated BDNF mRNA and in particular NT‐4 mRNA. By contrast, TrkB mRNA remained constant at ages or time points at which other mRNAs amplified from the very same cDNA libraries revealed dramatic increases or decreases. Our study suggests the existence of a complex regulatory neurotrophin network controlling the expression of other neurotrophins.
Voltage‐gated K+ current: a marker for apoptosis in differentiating neuronal progenitor cells?
Authors: Hribar Marusa , Bloc Alain , Medilanski Jeana , Nüsch Lars , Eder‐Colli Lorenza
Published: July 2004

We investigated apoptosis during early stages of in vitro differentiation of neuronal precursors generated by embryonic day 14 (E14) mouse striata stem cells. Differentiation was in conditions of suboptimal growth factor supply. Apoptosis reached 10–15% of cells and affected proliferating as well as postmitotic cells, including TUJ1‐positive cells. Inhibition of apoptosis led to an increased proportion of TUJ1‐positive cells generated by stem cells. K+ current was reported to be related to apoptosis. Outward K+ currents were present in differentiating neuronal precursors that were consistent with delayed rectifier and transient A‐type currents. The amplitude of the delayed rectifier current varied during the first 4 days of stem cell differentiation. Current amplitude was greatly increased in the presence of staurosporine but reduced at elevated extracellular K+ concentration. In addition, the amplitude of the current was significantly diminished by inhibiting several caspases, but not caspase 8. In Bax knock‐out transgenic neuronal precurors, K+ current was not decreased after the first day but at later stages of cell differentiation. At this early stage, apoptosis of proliferating cells and of TUJ1‐positive cells was not reduced by the absence of Bax, but was by caspase 9 inhibition. Thus, activation of a delayed rectifier K+ current in differentiating stem cells is related to apoptosis. Recordings of this current revealed that apoptosis at early stages of neuronal differentiation occurred in two phases that did not exhibit similar dependence on the proapoptotic protein Bax and that probably used different pathways.
Brain‐derived neurotrophic factor is expressed in a gradient in the superior colliculus during development of the retinocollicular projection
Authors: Marotte L. R. , Vidovic M. , Wheeler E. , Jhaveri S.
Published: July 2004

Theoretical models of topographic map formation have postulated a gradient of attractant in addition to a gradient of repulsion in the target. In species where many axons grow past their correct positions initially, it has also been argued that a parallel gradient of attractant or branching signal is required to ensure collateral formation at the correct position (O'Leary et al., 1999). Brain‐derived neurotrophic factor (BDNF) is a known attractant and promotes branching of retinal axons. We have examined its distribution in the superior colliculus and that of its receptor, trkB, in the retina, using immunohistochemistry and in situ hybridization, respectively, during the development of the topographic retinocollicular projection in the wallaby, a marsupial mammal. The number of glial endfeet expressing BDNF at the surface of the colliculus was found to be in a high caudal‐to‐low rostral gradient during the time when the retinocollicular projection was developing. When the projection was mature the rostrocaudal gradient had disappeared and the number of detectable endfeet expressing BDNF was very low. Messenger RNA for TrkB was expressed in the retinal ganglion cell layer throughout the time when the retinocollicular projection was developing, with no difference in expression across the nasotemporal axis of the retina. The low rostral to high caudal distribution of BDNF in glial endfeet supports the idea that it is providing a parallel gradient of attractant or branching signal in the colliculus.
Reward expectation, orientation of attention and locus coeruleus‐medial frontal cortex interplay during learning
Authors: Bouret Sebastien , Sara Susan J.
Published: July 2004

Regulation of attention and promotion of behavioural flexibility are functions attributed to both the noradrenergic nucleus locus coeruleus (LC) and the prefrontal cortex (PFC). The PFC receives a large innervation from LC and small changes in catecholaminergic activity in PFC profoundly affect cognitive function. It is crucial to the understanding of learning‐related plasticity, that the cognitive context driving LC neurons be determined and the relation to activity in PFC be elucidated. To this end simultaneous recordings were made from LC and prelimbic cortex (PL) during an odour‐reward association task in the rat. Neuronal activity related to orientation of attention, reward predictability, reward itself, and changes in stimulus reinforcement contingencies, was measured. All LC neurons and a significant proportion of PL neurons were engaged during several aspects of a Go/NoGo task, especially after the signal for trial onset and CS+ presentation. LC activation was, however, more tightly aligned to the behavioural response than to the CS+ 22% of PL neurons were activated during the response‐reward delay. This suggests that the activity of both these structures is related to reward anticipation. Finally, LC neurons exhibited rapid plasticity when the reward‐contingency was modified. Within‐trial response latencies were always shorter in LC than in PL and between‐trial response adaptation in LC preceded that in PL by many trials. Identifying such temporal relationships is an essential step toward understanding how neuromodulatory inputs to forebrain networks might promote or permit experience‐dependent plasticity in behavioural situations.
Flies lacking all synapsins are unexpectedly healthy but are impaired in complex behaviour
Authors: Godenschwege Tanja A. , Reisch Dietmar , Diegelmann Sören , Eberle Kai , Funk Natalja , Heisenberg Martin , Hoppe Viviane , Hoppe Jürgen , Klagges Bert R. E. , Martin Jean‐René , Nikitina Ekaterina A. , Putz Gabi , Reifegerste Rita , Reisch Natascha , Rister Jens , Schaupp Michael , Scholz Henrike , Schwärzel Martin , Werner Ursula , Zars Troy D. , Buchner Sigrid , Buchner Erich
Published: July 2004

Vertebrate synapsins are abundant synaptic vesicle phosphoproteins that have been proposed to fine‐regulate neurotransmitter release by phosphorylation‐dependent control of synaptic vesicle motility. However, the consequences of a total lack of all synapsin isoforms due to a knock‐out of all three mouse synapsin genes have not yet been investigated. In Drosophila a single synapsin gene encodes several isoforms and is expressed in most synaptic terminals. Thus the targeted deletion of the synapsin gene of Drosophila eliminates the possibility of functional knock‐out complementation by other isoforms. Unexpectedly, synapsin null mutant flies show no obvious defects in brain morphology, and no striking qualitative changes in behaviour are observed. Ultrastructural analysis of an identified ‘model’ synapse of the larval nerve muscle preparation revealed no difference between wild‐type and mutant, and spontaneous or evoked excitatory junction potentials at this synapse were normal up to a stimulus frequency of 5 Hz. However, when several behavioural responses were analysed quantitatively, specific differences between mutant and wild‐type flies are noted. Adult locomotor activity, optomotor responses at high pattern velocities, wing beat frequency, and visual pattern preference are modified. Synapsin mutant flies show faster habituation of an olfactory jump response, enhanced ethanol tolerance, and significant defects in learning and memory as measured using three different paradigms. Larval behavioural defects are described in a separate paper. We conclude that Drosophila synapsins play a significant role in nervous system function, which is subtle at the cellular level but manifests itself in complex behaviour.
Axonal injury‐dependent induction of the peripheral benzodiazepine receptor in small‐diameter adult rat primary sensory neurons
Authors: Karchewski Laurie A. , Bloechlinger Stefan , Woolf Clifford J.
Published: July 2004

The peripheral benzodiazepine receptor (PBR), a benzodiazepine but not γ‐aminobutyric acid‐binding mitochondrial membrane protein, has roles in steroid production, energy metabolism, cell survival and growth. PBR expression in the nervous system has been reported in non‐neuronal glial and immune cells. We now show expression of both PBR mRNA and protein, and the appearance of binding of a synthetic ligand, [3H]PK11195, in dorsal root ganglion (DRG) neurons following injury to the sciatic nerve. In naïve animals, PBR mRNA, protein expression and ligand binding are undetectable in the DRG. Three days after sciatic nerve transection, however, PBR mRNA begins to be expressed in injured neurons, and 4 weeks after the injury, expression and ligand binding are present in 35% of L4 DRG neurons. PBR ligand binding also appears after injury in the superficial dorsal horn of the spinal cord. The PBR expression in the DRG is restricted to small and medium‐sized neurons and returns to naïve levels if the injured peripheral axons are allowed to regrow and reinnervate targets. No non‐neuronal PBR expression is detected, unlike its putative endogenous ligand the diazepam binding inhibitor (DBI), which is expressed only in non‐neuronal cells, including the satellite cells that surround DRG neurons. DBI expression does not change with sciatic nerve transection. PBR acting on small‐calibre neurons could play a role in the adaptive survival and growth responses of these cells to injury of their axons.
Latent inhibition of cued fear conditioning: an NMDA receptor‐dependent process that can be established in the presence of anisomycin
Authors: Lewis Michael C. , Gould Thomas J.
Published: July 2004

Much of the research examining the biological basis for long‐term memories has focused on mechanisms that support the formation of conditioned associations. Less information is available on biological mechanisms which underlie processes that modify the strength of conditioned associations. Latent inhibition is a phenomenon by which pre‐exposure to a to‐be‐conditioned stimulus (CS) weakens subsequent conditioning of that CS to an unconditioned stimulus (US). Here we report that latent inhibition of cued fear conditioning is dependent on NMDA receptor activation. MK‐801 (1 mg/kg), an NMDA receptor antagonist, abolished latent inhibition of cued fear conditioning. This dose of MK‐801 administered before training did not disrupt cued fear conditioning. Conversely, anisomycin (150 mg/kg), a protein synthesis inhibitor, had no effect on latent inhibition of cued fear conditioning when administered 20 min before, immediately after, or 2, 4, 6, or 8 h after CS pre‐exposure. Furthermore, continuous anisomycin administration (50 mg/kg, administered every 2 h for 6 h starting 20 min prior to pre‐exposure) did not disrupt latent inhibition of cued fear conditioning. In addition, anisomycin had no effect on a long‐lasting version of latent inhibition of cued fear conditioning that was maintained over a 7‐day interval. Anisomycin administered before training, however, disrupted learning of the CS–US association. These findings suggest that latent inhibition of cued fear conditioning is a long‐lasting NMDA receptor‐dependent process that can develop during the inhibition of protein synthesis.
Functional topology of the mossy fibre–granule cell– Purkinje cell system revealed by imaging of intrinsic fluorescence in mouse cerebellum
Authors: Coutinho V. , Mutoh H. , Knöpfel T.
Published: July 2004

We report an activity‐induced green fluorescence signal observed when mouse cerebellar slices were illuminated with blue light and parallel fibre–Purkinje cell synapses were activated. The optical signal consisted of an initial increase in fluorescence that peaked within 1–2 s after the onset of stimulation, followed by a long lasting (40 s) transient decrease in fluorescence. Single or tetanic electrical stimuli applied to the molecular layer elicited ‘beam‐shaped’ fluorescence changes along the trajectory of parallel fibres. These signals reported activation of Purkinje cells as they were depressed by antagonists of ionotropic and metabotropic glutamate receptors at Purkinje cells and correlated with Purkinje cell spiking activity. Optical responses induced by direct pharmacological activation of glutamate receptors were reduced by a calcium‐free extracellular medium, consistent with the hypothesis that they reflect metabolic activity due to an increased intracellular calcium load associated with neuronal activation. We used these intrinsic fluorescence signals to address the question of whether granule cells excite Purkinje cells only locally via the ascending branches of their axons, or more widespread along the parallel fibre trajectory. White matter stimulation of the mossy fibres also elicited a beam‐like fluorescence change along the trajectory of parallel fibres. Simultaneous imaging and extracellular recording demonstrated the association between the beam‐like fluorescence signal and Purkinje cell spiking. This non‐invasive imaging technique supports the notion that parallel fibre activity, evoked either locally or through the mossy fibre–granule cell pathway, can activate postsynaptic Purkinje cells along more than 3 mm of the parallel fibre trajectory.
Restricted diffusion in the brain of transgenic mice with cerebral amyloidosis
Authors: Mueggler Thomas , Meyer‐Luehmann Melanie , Rausch Martin , Staufenbiel Matthias , Jucker Mathias , Rudin Markus
Published: July 2004

A prominent hallmark of Alzheimer's disease pathology is cerebral amyloidosis. However, it is not clear how extracellular amyloid‐β peptide (Aβ) deposition and amyloid formation compromise brain function and lead to dementia. It has been argued that extracellular amyloid deposition is neurotoxic and/or that soluble Aβ oligomers impair synaptic function. Amyloid deposits, by contrast, may affect diffusion properties of the brain interstitium with implications for the transport of endogenous signalling molecules during synaptic and/or extrasynaptic transmission. We have used diffusion‐weighted magnetic resonance imaging to study diffusion properties in brains of young (6‐month‐old) and aged (25‐month‐old) APP23 transgenic mice and control littermates. Our results demonstrate that fibrillar amyloid deposits and associated gliosis in brains of aged APP23 transgenic mice are accompanied by a reduction in the apparent diffusion coefficient. This decrease was most pronounced in neocortical areas with a high percentage of congophilic amyloid and was not significant in the caudate putamen, an area with only modest and diffuse amyloid deposition. These findings suggest that extracellular deposition of fibrillar amyloid and/or associated glial proliferation and hypertrophy cause restrictions to interstitial fluid diffusion. Reduced diffusivity within the interstitial space may alter volume transmission and therefore contribute to the cognitive impairment in Alzheimer's disease.
Time‐dependent changes in the expression of the MEF2 transcription factor family during topographic map reorganization in mammalian visual cortex
Authors: Leysen Inge , Van der Gucht Estel , Eysel Ulf T. , Huybrechts Roger , Vandesande Frans , Arckens Lutgarde
Published: July 2004

Removal of retinal input from a restricted region of adult mammalian visual cortex leads to a substantial reorganization of the retinotopy within the lesion projection zone (LPZ) of primary visual cortex (area 17). Little is known about the molecular mechanisms underlying such cortical plasticity. We investigated whether small but homonymous central retinal lesions induced differences in gene expression patterns between central area 17, the LPZ, vs. peripheral area 17 of the adult cat. Systematic differential mRNA display screening revealed higher levels for the mRNA encoding the transcription factor MEF2A in the LPZ. Semi‐quantitative PCR confirmed this dependency of mef2A mRNA expression on visual eccentricity in area 17 of animals with retinal lesions in contrast to normal animals. Western blotting experiments extended these data to the protein level and to two other members of the MEF2 transcription factor family, i.e. MEF2C and MEF2D. Quantitative analysis of the Western blotting experiments further revealed a post‐lesion survival time‐dependent change in expression for all three MEF2 family members. The lesion effect was maximal at 3 days and 1 month post‐lesion, but only minor at 2 weeks post‐lesion. Interestingly, complete removal of retinal input from area 17 by surgery did not significantly alter the expression of the MEF2 transcription factors, excluding a definite correlation between neuronal activity and MEF2A expression levels. MEF2A immunocytochemistry confirmed both qualitatively and quantitatively the Western blotting observations in all animal models. Together, our findings identified a brain plasticity‐related expression pattern for the MEF2 transcription factor family in adult mammalian neocortex.