bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of Potential Antagonists of Human Interleukin 5 Demonstrates Their Cross‐Reactivity with Receptors for Interleukin 3 and Granulocyte‐Macrophage Colony‐Stimulating Factor
Authors: Wiekowski Maria , Prosser Dina , Taremi Shane , Tsarbopoulos Anthony , Jenh Chung‐Her , Chou Chuan‐Chu , Lundell Dan , Zavodny Paul , Narula Satwant
Published: July 2004

The ligand‐binding α‐chain of the human interleukin 5 (IL‐5) receptor was expressed in its soluble form, lacking the transmembrane and cytoplasmic domains, from recombinant baculoviras. The soluble receptor was used in a scintillation proximity assay to identify two chemical compounds that inhibit binding of human IL‐5 to the soluble receptor α chain with IC50 of 8 μM and 11 μM. These compounds also inhibited the interaction of human IL‐5 with its membrane‐bound receptor, composed of the ligand‐binding a chain and signal‐transducing β chain, and prevented signaling through the receptor. Analysis by surface plasmon resonance and matrix‐assisted laser‐desorption/ionization mass spectrometry showed that the identified compounds bound irreversibly to the receptor at a 1:1 (mol/mol) ratio, suggesting a covalent interaction with the α chain of the human IL‐5 receptor. Both compounds also inhibited the interaction of the receptors for interleukin 3 (IL‐3) and granulocyte‐macrophage colony‐stimulating factor (GM‐CSF), which are involved in hematopoietic differentiation and activation of immune cells, thus eliminating them as potential therapeutic agents. The inhibition of the structurally closely related receptors for IL‐5, IL‐3 and GM‐CSF by both compounds, while binding of interleukin‐4 to its receptor was not affected, suggests that a similar reactive site exists in the ligand‐binding domains of the receptors for IL‐5, IL‐3 and GM‐CSF.
The Effect of α‐Tocopherol on the Synthesis, Phosphorylation and Activity of Protein Kinase C in Smooth Muscle Cells After Phorbol 12‐Myristate 13‐Acetate Down‐Regulation
Authors: Clément Sophie , Tasinato Andrea , Boscoboinik Daniel , Azzi Angelo
Published: July 2004

Previous work had established that, in smooth muscle cells, α‐tocopherol negatively regulates protein kinase C by reventing its activation [Tasinato, A., Boscoboinik, D., Bartoli, G. M., Maroni, P. & Azzi, A. (1995)Proc. Natl Acad. Sci. USA 92, 12190–12194]. In this study, the mechanism by which this event takes place has been analyzed. The regulation by α‐tocopherol of protein kinase C expression, activity and phosphorylation has been followed during the synthesis of protein kinase C after its down‐regulation by phorbol 12‐myristate 13‐acetate. The data show that protein kinase C isoenzyme α is synthesised significantly more (30% 72 h after down‐regulation) in the presence of α‐tocopherol. However, its activity is significantly less (45% diminution) and its phosphorylation state is also decreased (60% diminution). The effect of α‐tocopherol appears not to be shared by the analogue β‐tocopherol, provided with similar radical‐scavenging properties. The data are interpreted in terms of a diminution of protein kinase C phosphorylation, specifically caused by α‐tocopherol, resulting in a decreased enzyme specific activity.
C‐Terminal Extension of the H2‐Activating Subunit, HoxH, Directs Maturation of the NAD‐Reducing Hydrogenase in Alcaligenes Eutrophus
Authors: Massanz Christian , Fernandezz Victor M. , Friedrich Bärbel
Published: July 2004

Formation of enzymatically active [NiFe] hydrogenases is dependent on a number of posttranslational steps, including metal attachment to a precursor of the catalytic subunit, truncation of a small C‐terminal peptide from the precursor, and oligomerisation of the subunits. Two amino acid replacements were introduced by site‐directed mutagenesis at the C‐terminal proteolytic cleavage site of HoxH, the Ni‐containing subunit of the cytoplasmic NAD‐reducing hydrogenase of Alcaligenes eutrophus H16. Replacement of Ala465, the first residue of the 24‐amino‐acid cleaved polypeptide, by Pro yielded a form of HoxH that was blocked in C‐terminal proteolysis. This HoxH subunit, although capable of binding Ni, was blocked in formation of a stable tetrameric holoenzyme. In the second mutant, the C‐terminal extension of HoxH was eliminated by substituting the Ala codon for a translational stop codon. Although this mutant subunit was able to form the oligomeric holoenzyme, it was devoid of Ni. Both mutant proteins contained only traces of H2‐activating functions. H2‐dependent reduction of NAD and benzylviologen, and D2/H+‐exchange activity were almost completely abolished, while the NADH oxidoreductase activity, mediated by the diaphorase moiety of the hydrogenase, was retained. These results allow the following conclusions: the C‐terminal extension of HoxH is neccessary to direct specific Ni insertion into the hydrogenase; subunit assembly to the holoenzyme is not dependent on Ni insertion; and a precursor with the C‐terminal peptide is not competent for assembly.
N‐Terminal segment of potato virus X coat protein subunits is glycosylated and mediates formation of a bound water shell on the virion surface
Authors: Baratova Lyudmila A. , Fedorova Nataliya V. , Dobrov Eugenie N. , Lukashina Elena V. , Kharlanov Andrey N. , Nasonov Vitaly V. , Serebryakova Marina V. , Kozlovsky Stanislav V. , Zayakina Olga V. , Rodionova Nina P.
Published: July 2004

The primary structures of N‐terminal 19‐mer peptides, released by limited trypsin treatment of coat protein (CP) subunits in intact virions of three potato virus X (PVX) isolates, were analyzed. Two wild‐type PVX strains, Russian (Ru) and British (UK3), were used and also the ST mutant of UK3 in which all 12 serine and threonine residues in the CP N‐terminal segment were replaced by glycine or alanine. With the help of direct carbohydrate analysis and MS, it was found that the acetylated N‐terminal peptides of both wild‐type strains are glycosylated by a single monosaccharide residue (galactose or fucose) at NAcSer in the first position of the CP sequence, whereas the acetylated N‐terminal segment of the ST mutant CP is unglycosylated. Fourier transform infrared spectra in the 1000–4000 cm−1 region were measured for films of the intact and in situ trypsin‐degraded PVX preparations at low and high humidity. These spectra revealed the presence of a broad‐band in the region of valent vibrations of OH bonds (3100–3700 cm−1), which can be represented by superposition of three bands corresponding to tightly bound, weakly bound, and free OH groups. On calculating difference (‘wet’ minus ‘dry’) spectra, it was found that the intact wild‐type PVX virions are characterized by high water‐absorbing capacity and the ability to order a large number of water molecules on the virus particle. This effect was much weaker for the ST mutant and completely absent in the trypsin‐treated PVX. It is proposed that the surface‐located and glycosylated N‐terminal CP segments of intact PVX virions induce the formation of a columnar‐type shell from bound water molecules around the virions, which probably play a major role in maintaining the virion surface structure.
Mode of action of the microbial metabolite GE23077, a novel potent and selective inhibitor of bacterial RNA polymerase
Authors: Sarubbi Edoardo , Monti Federica , Corti Emiliana , Miele Anna , Selva Enrico
Published: July 2004

GE23077, a novel microbial metabolite recently isolated from Actinomadura sp. culture media, is a potent and selective inhibitor of bacterial RNA polymerase (RNAP). It inhibits Gram‐positive (Bacillus subtilis) and Gram‐negative (Escherichia coli) RNAPs with IC50 values (i.e. the concentration at which the enzyme activity is inhibited by 50%) in the 10−8 m range, whereas it is not active on E. coli DNA polymerase or on eukaryotic (wheat germ) RNAP II (IC50 values > 10−4 m in both cases). In spite of its potent activity on purified bacterial RNAPs, GE23077 shows a narrow spectrum of antimicrobial activity on Gram‐positive and Gram‐negative bacteria. To investigate the molecular basis of this behaviour, the effects of GE23077 on macromolecular biosynthesis were tested in E. coli cells permeabilized under different conditions. The addition of GE23077 to plasmolyzed cells resulted in an immediate and specific inhibition of intracellular RNA biosynthesis, in a dose–response manner, strongly suggesting that cell penetration is the main obstacle for effective antimicrobial activity of the antibiotic. Biochemical studies were also conducted with purified enzymes to obtain further insights into the mode of action of GE23077. Interestingly, the compound displays a behaviour similar to that of rifampicin, an antibiotic structurally unrelated to GE23077: both compounds act at the level of transcription initiation, but not on the σ subunit and not on the formation of the promoter DNA–RNAP complex. Tests on different rifampicin‐resistant E. coli RNAPs did not show any cross‐resistance between the two compounds, indicating distinct binding sites on the target enzyme. In conclusion, GE23077 is an interesting new molecule for future mechanistic studies on bacterial RNAP and for its potential in anti‐infective drug discovery.
Protein transport into canine pancreatic microsomes
Authors: Guth Silvia , Völzing Christian , Müller Anika , Jung Martin , Zimmermann Richard
Published: July 2004

Transport of presecretory proteins into the mammalian rough endoplasmic reticulum involves a protein translocase that comprises the integral membrane proteins Sec61αp, Sec61βp, and Sec61γp as core components. Electron microscopic analysis of protein translocase in rough microsomal membranes suggested that between three and four heterotrimeric Sec61p complexes form the central unit of protein translocase. Here we analyzed the stoichiometry of heterotrimeric Sec61p complexes present in cotranslationally active protein translocases of canine pancreatic microsomes and various other lumenal and membrane components believed to be subunits of protein translocase and to be involved in covalent modifications. Based on these numbers, the capacity for cotranslational transport was estimated for the endoplasmic reticulum of the human pancreas.
Expression, purification and catalytic activity of Lupinus luteus asparagine β‐amidohydrolase and its Escherichia coli homolog
Authors: Borek Dominika , Michalska Karolina , Brzezinski Krzysztof , Kisiel Agnieszka , Podkowinski Jan , Bonthron David T. , Krowarsch Daniel , Otlewski Jacek , Jaskolski Mariusz
Published: July 2004

We describe the expression, purification, and biochemical characterization of two homologous enzymes, with amidohydrolase activities, of plant (Lupinus luteus potassium‐independent asparaginase, LlA) and bacterial (Escherichia coli, ybiK/spt/iaaA gene product, EcAIII) origin. Both enzymes were expressed in E. coli cells, with (LlA) or without (EcAIII) a His‐tag sequence. The proteins were purified, yielding 6 or 30 mg·L−1 of culture, respectively. The enzymes are heat‐stable up to 60 °C and show both isoaspartyl dipeptidase and l‐asparaginase activities. Kinetic parameters for both enzymatic reactions have been determined, showing that the isoaspartyl peptidase activity is the dominating one. Despite sequence similarity to aspartylglucosaminidases, no aspartylglucosaminidase activity could be detected. Phylogenetic analysis demonstrated the relationship of these proteins to other asparaginases and aspartylglucosaminidases and suggested their classification as N‐terminal nucleophile hydrolases. This is consistent with the observed autocatalytic breakdown of the immature proteins into two subunits, with liberation of an N‐terminal threonine as a potential catalytic residue.
Arginine ethylester prevents thermal inactivation and aggregation of lysozyme
Authors: Shiraki Kentaro , Kudou Motonori , Nishikori Shingo , Kitagawa Harue , Imanaka Tadayuki , Takagi Masahiro
Published: July 2004

Arginine is a versatile additive to prevent protein aggregation. This paper shows that arginine ethylester (ArgEE) prevents heat‐induced inactivation and aggregation of hen egg lysozyme more effectively than arginine or guanidine. The addition of ArgEE decreased the melting temperature of lysozyme. This data could be interpreted in terms of ArgEE binding to unfolded lysozyme, possibly through the ethylated carboxyl group, which leads to effective prevention of intermolecular interaction among aggregation‐prone molecules. The data suggest that ArgEE could be used as an additive to prevent inactivation and aggregation of heat‐labile proteins.
Growth of Differentiated Ovine Tracheal Epithelial Cells In Vitro
Authors: Radi Z. A. , Ackermann M. R.
Published: July 2004

Culture of ovine tracheal epithelial cells is a useful tool for conducting various in vitro studies. We describe herein an in vitro technique and the conditions for culturing primary epithelial cells derived from tracheas of adult sheep. Ovine tracheas were surgically removed from 2‐ to 3‐month‐old healthy sheep and tracheal epithelial cells were isolated by 0.15% pronase digestion. After epithelial cells isolation, a Millicell insert with porous membrane was coated with 0.05% human placental collagen and the epithelial cells were added to the membrane. To create an air–liquid interface environment for the cells, the apical compartment of the membrane containing the tracheal epithelial cells was left exposed to 5% CO2 at 37°C for 2 days then increased to 9% CO2 while cells in the basolateral compartment underneath the membrane contained the growth medium necessary for cells nourishment. Pepsin digestion was more effective in reducing the number of fibroblasts than other procedures. Cells were allowed to grow for 6–7 days to form a confluent monolayer and nearly 21 days for cilia formation on the apical surface as determined by light microscopy of haematoxylin and eosin‐stained sections of membranes. In order to further confirm the epithelial origin of cells, cells were stained for cytokeratin antigen by immunohistochemistry. Most ciliated epithelial cells were immunoreactive for cytokeratin. This is the first report of differentiated ovine tracheal epithelial cells growth and isolation. This technique can be used in numerous in vitro investigative studies in ovine species as an animal model for human disease.
The Effect of Parentage on the Prevalence, Severity and Location of Lesions of Osteochondrosis in Swine
Authors: Ytrehus B. , Grindflek E. , Teige J. , Stubsjøen E. , Grøndalen T. , Carlson C. S. , Ekman S.
Published: July 2004

The aims of this study were to determine the effects of parentage and gender on the prevalence, severity and location of lesions of osteochondrosis manifesta (OCM) and osteochondrosis dissecans (OCD) in offspring from different Norwegian Landrace boars and to examine the relationships between lesion characteristics and selected growth parameters. Fifteen sires were selected based on their high breeding value for osteochondrosis. Seven locations in the distal humerus and the distal femur from 1680 offspring of these animals were evaluated for severity of OCM and presence of OCD by gross examination of serially sectioned humeri and femora. Osteochondrosis manifesta was most prevalent in the trochlea of humerus, the sagittal ridge of humerus, the medial condyle of femur and the medial sulcus obliquus of femur. The severity of the lesions and the prevalence of OCD were highest in the trochlea and the sagittal ridge of humerus. Castrates had significantly higher OCM scores than sows. There were significant effects of both sire and dam on the OCM scores of the offspring in most locations; however, growth rate and weight at slaughter did not influence the OCM score.
Evaluation of Stratification Factors and Score‐scales in Clinical Trials of Treatment of Clinical Mastitis in Dairy Cows
Authors: Hektoen L. , Ødegaard S. A. , Løken T. , Larsen S.
Published: July 2004

There is often a need to reduce sample size in clinical trials due to practical limitations and ethical considerations. Better comparability between treatment groups by use of stratification in the design, and use of continuous outcome variables in the evaluation of treatment results, are two methods that can be used in order to achieve this. In this paper the choice of stratification factors in trials of clinical mastitis in dairy cows is investigated, and two score‐scales for evaluation of clinical mastitis are introduced. The outcome in 57 dairy cows suffering from clinical mastitis and included in a clinical trial comparing homeopathic treatment, placebo and a standard antibiotic treatment is investigated. The strata of various stratification factors are compared across treatments to determine which other factors influence outcome. The two score scales, measuring acute and chronic mastitis symptoms, respectively, are evaluated on their ability to differentiate between patients classified from clinical criteria as responders or non‐responders to treatment. Differences were found between the strata of the factors severity of mastitis, lactation number, previous mastitis this lactation and bacteriological findings. These factors influence outcome of treatment and appear relevant as stratification factors in mastitis trials. Both score scales differentiated between responders and non‐responders to treatment and were found useful for evaluation of mastitis and mastitis treatment.
Effect of Vitamin E Supplementation on Various Functional Properties of Macrophages and Neutrophils Obtained from Weaned Piglets
Authors: Fragou S. , Fegeros K. , Xylouri E. , Baldi A. , Politis I.
Published: July 2004

Sixteen piglets were used to determine the effect of vitamin E supplementation on several functional properties of macrophages and neutrophils obtained from weaned piglets. Piglets, immediately following weaning, were assigned to one of three experimental groups: control (no vitamin E supplementation), low level of vitamin E supplementation (100 mgdl‐α‐tocopheryl acetate/kg diet) and high level of vitamin E supplementation (300 mgdl‐α‐tocopheryl acetate/kg diet). Supplementation of vitamin E lasted for a period of 36 days, following a 3‐day adaptation period after weaning. Blood samples were collected on days 0, 12, 24 and 36 of the experimental period, monocytes/macrophages and neutrophils were isolated and the following parameters were determined in macrophages and neutrophils activated by phorbol myristate acetate: total cell‐associated and membrane‐bound urokinase plasminogen activator (u‐PA) activity and superoxide anion production. Results showed that macrophages and neutrophils isolated from piglets that received supplemental vitamin E had higher (P < 0.05) total and membrane‐bound u‐PA activities as well as higher (P < 0.05) superoxide anion production compared with the values of the corresponding cells obtained from control piglets on day 12 of the experimental period. Both levels of vitamin E supplementation (low and high) were equally effective. In contrast, vitamin E supplementation had no effect (P > 0.05) on total and membrane‐bound u‐PA activities and superoxide anion production by porcine macrophages and neutrophils on days 24 and 36 of the experimental period. In conclusion, the low level of vitamin E supplementation is recommended for piglets for the first 2 weeks after weaning.
Comparison of the Effects of Fructose and Glucose Supplementation on Metabolic Responses in Resting and Exercising Horses
Authors: Vervuert I. , Coenen M. , Bichmann M.
Published: July 2004

The aim of this study was to compare the effects of two different carbohydrate sources (fructose and glucose) on the metabolic responses in resting and exercising horses. The following regimes were fed in randomized order to five trained horses at rest and immediately before or during exercise. The resting regime comprised 0.6 kg grass meal pellets (control) or 0.6 kg grass meal pellets supplemented with either 50% glucose or 50% fructose. The exercise regime comprised 0.3 kg grass meal (control) or 0.6 kg grass meal pellets supplemented with either 50% glucose or 50% fructose fed immediately before or during simulated endurance exercise on a treadmill (30 km, total running time 120 min; 15 min rest after 60 min running time). Blood samples were collected for the analysis of glucose, insulin, free fatty acids (FFA) and lactate. In resting horses, glucose supplementation resulted in a significantly higher glycaemic and insulinaemic response than the control or fructose feedings (treatmentP < 0.05). Plasma glucose levels were significantly higher after glucose supplementation given before or during exercise. Similar plasma glucose concentrations were observed after fructose was fed before exercise, whereas fructose supplementation during exercise resulted in a lower glycaemic response than glucose feeding (P < 0.05). Plasma insulin, FFA and lactate levels showed exercise‐related changes (timeP < 0.05), but treatment did not effect these results. Plasma glucose concentrations were elevated during the simulated endurance exercise after glucose feeding, and no counter‐regulation by insulin occurred. The higher glycaemic response might be beneficial as exogenous glucose can be used as an energetic substrate during prolonged exercise. Fructose exhibited no clear advantages in comparison with glucose as a carbohydrate source for exercising horses.
Reduced ocular dominance plasticity and long‐term potentiation in the developing visual cortex of protein kinase A RIIα mutant mice
Authors: Rao Yan , Fischer Quentin S. , Yang Yupeng , McKnight G. Stanley , LaRue Adrienne , Daw Nigel W.
Published: July 2004

The cAMP‐dependent protein kinase (PKA) signalling pathway has been shown to play an important role in long‐term potentiation (LTP) and depression (LTD), and ocular dominance plasticity in the visual cortex. In order to investigate further the involvement of individual PKA subunits in visual cortical plasticity, LTP and LTD in vitro and ocular dominance plasticity in vivo in the developing visual cortex were examined in mice lacking the RIIα subunit of PKA. Here we show that LTP in layers II/III was decreased in RIIα knockout mice, but LTD was almost unaffected, and the ocular dominance shift induced by monocular deprivation was also partially blocked. These data provide evidence that RIIα is involved in LTP and ocular dominance plasticity, and further suggest that different afferent inputs could selectively activate particular subunits of PKA and thereby direct specific aspects of visual cortical plasticity.
Blockade of presynaptic adenosine A1 receptor responses by nitric oxide and superoxide in rat hippocampus
Authors: Shahraki Ali , Stone Trevor W.
Published: July 2004

Activation ofN‐methyl‐d‐aspartate (NMDA) receptors prevents the neuronal responses to adenosine in hippocampal slices. As NMDA receptor activation leads to the generation of nitric oxide (NO) and superoxide, we have examined whether these can modify neuronal responses to adenosine and mediate the actions of NMDA. Field excitatory postsynaptic potentials were recorded in the CA1 region of rat hippocampal slices. Paired‐pulse interactions were studied to localize the observed interactions to presynaptic terminals. The NO donorsS‐nitroso‐N‐acetylpenicillamine and diethylamine NONOate induced a long‐lasting potentiation (NO‐induced potentiation) of field excitatory postsynaptic potential slope and significantly prevented the presynaptic inhibitory effect of adenosine or the A1 receptor agonist N6‐cyclopentyladenosine selectively with no effect on responses to baclofen. The superoxide‐generating system of xanthine/xanthine oxidase also prevented presynaptic responses to adenosine and this effect was prevented by superoxide dismutase (SOD). The guanylate cyclase inhibitor 1H‐[1,2,4]‐oxadiazolo[4,3a]quinoxalin‐1‐one (10 µm) prevented NO‐induced potentiation and the inhibitory effects ofS‐nitroso‐N‐acetylpenicillamine and xanthine/xanthine oxidase on adenosine responses. The inhibitory effect of NMDA on adenosine responses was unchanged by 1H‐[1,2,4]‐oxadiazolo[4,3a]quinoxalin‐1‐one, indicating that guanosine‐3′,5‐cyclic monophosphate does not mediate this interaction, although it was partially reduced by SOD, suggesting that superoxide might contribute. The reduction of adenosine responses by electrically‐induced long‐term potentiation was prevented by NO synthase inhibition or SOD. The results indicate that the presynaptic effects of adenosine at presynaptic sites can be prevented by NO or superoxide but that neither of these individually can fully account for the prevention of adenosine responses by NMDA.
Trophic support delays but does not prevent cell‐intrinsic degeneration of neurons deficient for munc18‐1
Authors: Heeroma Joost H. , Roelandse Martijn , Wierda Keimpe , Van Aerde Karlijn I. , Toonen Ruud F. G. , Hensbroek Robert A. , Brussaard Arjen , Matus Andrew , Verhage Matthijs
Published: July 2004

The stability of neuronal networks is thought to depend on synaptic transmission which provides activity‐dependent maintenance signals for both synapses and neurons. Here, we tested the relationship between presynaptic secretion and neuronal maintenance using munc18‐1‐null mutant mice as a model. These mutants have a specific defect in secretion from synaptic and large dense‐cored vesicles [Verhage et al. (2000), Science, 287, 864–869; Voets et al. (2001), Neuron, 31, 581–591]. Neuronal networks in these mutants develop normally up to synapse formation but eventually degenerate. The proposed relationship between secretion and neuronal maintenance was tested in low‐density and organotypic cultures and, in vivo, by conditional cell‐specific inactivation of the munc18‐1 gene. Dissociated munc18‐1‐deficient neurons died within 4 days in vitro (DIV). Application of trophic factors, insulin or BDNF delayed degeneration up to 7 DIV. In organotypic cultures, munc18‐1‐deficient neurons survived until 9 DIV. On glial feeders, these neurons survived up to 10 DIV and 14 DIV when insulin was applied. Co‐culturing dissociated mutant neurons with wild‐type neurons did not prolong survival beyond 4 DIV, but coculturing mutant slices with wild‐type slices prolonged survival up to 19 DIV. Cell‐specific deletion of munc18‐1 expression in cerebellar Purkinje cells in vivo resulted in the specific loss of these neurons without affecting connected or surrounding neurons. Together, these data allow three conclusions. First, the lack of synaptic activity cannot explain the degeneration in munc18‐1‐null mutants. Second, trophic support delays but cannot prevent degeneration. Third, a cell‐intrinsic yet unknown function of munc18‐1 is essential for prolonged survival.
Stimulation of N‐methyl‐d‐aspartate receptors, AMPA receptors or metabotropic glutamate receptors leads to rapid internalization of AMPA receptors in cultured nucleus accumbens neurons
Authors: Mangiavacchi Simona , Wolf Marina E.
Published: July 2004

In hippocampus and other regions, α‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionate (AMPA) receptors are inserted into synapses during long‐term potentiation and removed during long‐term depression. However, little is known about regulation of AMPA receptor trafficking in the nucleus accumbens (NAc), despite growing evidence that glutamate‐dependent forms of plasticity in the NAc contribute to drug addiction. Using postnatal rat NAc cultures and an immunocytochemical method that selectively detects newly internalized GluR1, we studied the regulation of AMPA receptor internalization in NAc neurons by glutamate agonists. Newly internalized GluR1 was detected during 15 or 30 min of incubation at room temperature, indicating a basal rate of GluR1 turnover. The rate of GluR1 internalization was increased by glutamate (50 µm) within 5 min of its addition. Glutamate‐induced GluR1 internalization was partially blocked by either an AMPA receptor antagonist (CNQX; 20 µm) or an N‐methyl‐d‐aspartate (NMDA) receptor antagonist (APV; 50 µm). Both NMDA (50 µm) and AMPA (50 µm) increased GluR1 internalization in a Ca2+‐dependent manner. The NMDA effect was blocked by APV while the AMPA effect was blocked by APV or CNQX. We interpret these findings to suggest that NMDA and AMPA ultimately trigger GluR1 internalization through the same NMDA receptor‐dependent pathway. The effect of glutamate was also partially blocked by the group 1 metabotropic glutamate receptor antagonist N‐phenyl‐7‐(hydroxyimino)cyclopropa[b]chromen‐1a‐carboxamide (PHCCC; 50 µm), while the group 1 agonist 3,5‐dihydroxyphenylglycine (DHPG; 50 µm) stimulated GluR1 internalization. These data suggest that AMPA receptors on NAc neurons may be subject to rapid regulation of their surface expression in response to changes in the activity of glutamate inputs from cortical and limbic regions.
Anatomical and functional evidence for a role of arginine‐vasopressin (AVP) in rat olfactory epithelium cells
Authors: Levasseur Grégoire , Baly Christine , Grébert Denise , Durieux Didier , Salesse Roland , Caillol Monique
Published: July 2004

The olfactory epithelium (OE) is composed of olfactory sensory neurons (OSNs) and sustentacular cells; it lies in the nasal cavity where it is protected by a thin mucus layer. The finely regulated composition of this mucus provides OSN with a suitable ionic environment. To maintain the functional integrity of the epithelium despite permanent physical, chemical and microbial aggressions, both OSNs and surrounding sustentacular cells are continuously renewed from globose basal cells. Moreover, the sense of smell is involved in so numerous behaviours (feeding, reproduction, etc.) that it has to cross‐talk with the endocrine and neuroendocrine systems. Thus, besides its sensory function, the olfactory epithelium is thought to undergo a lot of complex regulatory processes. We therefore studied the effects of various neuropeptides on primary cultures of Sprague–Dawley rat olfactory epithelium cells. We found that arginine‐vasopressin (AVP) triggered a robust, dose‐dependent calcium increase in these cells. The cell response was essentially ascribed to the V1a AVP receptor, whose presence was confirmed by RT‐PCR and immunolabelling. In the culture, V1a but not V1b receptors were present, mainly localized in neurons. In the epithelium, both subtypes were found differentially distributed. V1a‐R were localized mainly in globose basal cells and at the apical side of the epithelium, in the area of the dendritic knobs of OSNs. V1b‐R were strongly associated with Bowman's gland cells and globose basal cells. These localizations suggested potential multifaceted roles of a hormone, AVP, in the olfactory epithelium.
Prolonged cannabinoid treatment results in spatial working memory deficits and impaired long‐term potentiation in the CA1 region of the hippocampus in vivo
Authors: Hill Matthew N. , Froc David J. , Fox Christopher J. , Gorzalka Boris B. , Christie Brian R.
Published: July 2004

Adult male Long‐Evans rats were administered the potent cannabinoid 1 receptor agonist HU‐210 (100 µg/kg, i.p.) for 15 days continuously and their performance on a matching‐to‐place version of the Morris water maze was subsequently evaluated. Overall, experimental animals performed significantly worse initially on the reference memory component of this task, but their performance improved over 5 days until it was indistinguishable from that of control animals. Animals given HU‐210 did not exhibit working memory impairments at short intertrial delays (30 s); however, significant impairments were observed in learning performance with longer intertrial delays (300 s). In vivo electrophysiological analyses revealed that long‐term potentiation in the CA1 region of the hippocampus was significantly impaired following the administration of HU‐210 for 15 days. These results indicate that long‐term cannabinoid exposure can produce marked deficits in reference and working memory performance, and also impair hippocampal synaptic plasticity in vivo.
Brain‐derived neurotrophic factor acutely depresses excitatory synaptic transmission to GABAergic neurons in visual cortical slices
Authors: Jiang Bin , Kitamura Akihiko , Yasuda Hiroki , Sohya Kazuhiro , Maruyama Atsushi , Yanagawa Yuchio , Obata Kunihiko , Tsumoto Tadaharu
Published: July 2004

Brain‐derived neurotrophic factor (BDNF) acutely modulates synaptic transmission to excitatory neurons in hippocampus and neocortex. The question of whether BDNF acts similarly on excitatory synaptic transmission to GABAergic neurons was eluded in previous studies using cortical slices. To address this question, we used transgenic mice in which expression of green fluorescence protein (GFP) is regulated by glutamic acid decarboxylase 67 (GAD67) promoter. In cortical slices prepared from these GAD67‐GFP knock‐in mice, we could detect GABAergic neurons under a fluorescent microscope. An application of BDNF rapidly depressed excitatory postsynaptic currents (EPSCs) evoked by layer IV stimulation in most GFP‐positive neurons in layer II/III of the cortex. This effect was seen at synapses activated during the BDNF application and blocked by anti‐TrkB IgG, indicating that the acute inhibitory action of BDNF is activity‐dependent and mediated through TrkB. Paired‐pulse ratios of the amplitude of EPSCs to paired stimulation at intervals of 10–100 ms were not significantly changed after BDNF application, suggesting that the site of depression may be postsynaptic. Responses to directly applied glutamate were also depressed by BDNF in most of neurons, being consistent with the interpretation of postsynaptic action of BDNF. The depressive action of BDNF was blocked by an intracellular injection of a Ca2+ chelator, suggesting that a rise in Ca2+ is involved in the acute depression of EPSCs. This action of BDNF was seen in 67% of parvalbumin (PV)‐positive neurons, but in only 19% of PV‐negative neurons, indicating that the depressive action is biased to PV‐positive GABAergic neurons.
In vivo31P NMR spectroscopy shows an increase in glycerophosphorylcholine concentration without alterations in mitochondrial function in the prefrontal cortex of medicated schizophrenic patients at rest
Authors: Shirayama Yukihiko , Yano Toshio , Takahashi Kiyohisa , Takahashi Seizo , Ogino Takashi
Published: July 2004

The 31P NMR localised method was used to study the metabolism of phospholipid and high energy phosphate in the prefrontal cortex. The spectra were taken from patients with schizophrenia (11 males) receiving neuroleptic medication, and were compared to normal controls (15 males). Their spectral intensities were analysed using a non‐linear least‐squares method with a prior knowledge of the fixed chemical shifts and linewidths, leading to further resolution into resonances of glycerophosphorylethanolamine (GPE), glycerophosphorylcholine (GPC), phosphorylethanolamine (PE) and phosphorylcholine (PC). The metabolite concentrations were calculated referring to the spectral intensities of phosphate phantoms with known concentrations. T1 values of phantom and cerebrum were estimated from a series of localised inversion recovery spectra to correct for the signal saturation effects. The schizophrenic patients showed an increased concentration of GPC but not GPE, PE or PC. Furthermore, no difference was observed regarding the concentration of high‐energy phosphates such as phosphocreatine, inorganic phosphate and ATP. The patients did not show any differences in mitochondrial function such as phosphorylation potential and the ratio of the rate of ATP synthesis. Thus, an increase in GPC concentration in the prefrontal cortex could be characteristic of the pathophysiology of schizophrenia with mild negative symptoms.
The rewarding properties of MDMA are preserved in mice lacking µ‐opioid receptors
Authors: Robledo Patricia , Mendizabal Victoria , Ortuño Jordi , De La Torre Rafael , Kieffer Brigitte L. , Maldonado Rafael
Published: July 2004

The involvement of µ‐opioid receptors in the rewarding properties of MDMA was explored in µ‐opioid receptor knockout mice using the conditioning place preference paradigm. The associated release of dopamine in the nucleus accumbens was investigated by in vivo microdialysis. A significant rewarding effect of MDMA (10 mg/kg, i.p.) was observed in both wild‐type and µ‐opioid receptor knockout mice. MDMA (10 mg/kg, i.p.) also induced similar increases in dopamine and decreases in 3,4‐dihydroxyphenylacetic acid and homovanillic acid in the nucleus accumbens dialysates of both wild‐type and µ‐opioid receptor knockout mice. No significant differences in basal levels of dopamine, 3,4‐dihydroxyphenylacetic or homovanillic acids between wild‐type and µ‐opioid receptor knockout mice were observed. In summary, the present results suggest that, in contrast to what has been reported for other drugs of abuse such as opioids, ethanol, nicotine and Δ9‐tetrahydrocannabinol, µ‐opioid receptors do not play a major role in the rewarding properties of MDMA. These differences could be due to distinct mechanisms controlling dopamine release in the nucleus accumbens and suggest that the effects of MDMA on dopaminergic neurons are independent of µ‐opioid receptors.
Kv3 K+ channels enable burst output in rat cerebellar Purkinje cells
Authors: McKay B. E. , Turner R. W.
Published: July 2004

The ability of cells to generate an appropriate spike output depends on a balance between membrane depolarizations and the repolarizing actions of K+ currents. The high‐voltage‐activated Kv3 class of K+ channels repolarizes Na+ spikes to maintain high frequencies of discharge. However, little is known of the ability for these K+ channels to shape Ca2+ spike discharge or their ability to regulate Ca2+ spike‐dependent burst output. Here we identify the role of Kv3 K+ channels in the regulation of Na+ and Ca2+ spike discharge, as well as burst output, using somatic and dendritic recordings in rat cerebellar Purkinje cells. Kv3 currents pharmacologically isolated in outside‐out somatic membrane patches accounted for ∼ 40% of the total K+ current, were very fast and high voltage activating, and required more than 1 s to fully inactivate. Kv3 currents were differentiated from other tetraethylammonium‐sensitive currents to establish their role in Purkinje cells under physiological conditions with current‐clamp recordings. Dual somatic‐dendritic recordings indicated that Kv3 channels repolarize Na+ and Ca2+ spikes, enabling high‐frequency discharge for both types of cell output. We further show that during burst output Kv3 channels act together with large‐conductance Ca2+‐activated K+ channels to ensure an effective coupling between Ca2+ and Na+ spike discharge by preventing Na+ spike inactivation. By contributing significantly to the repolarization of Na+ and especially Ca2+ spikes, our data reveal a novel function for Kv3 K+ channels in the maintenance of high‐frequency burst output for cerebellar Purkinje cells.
Decreased histamine H1 receptor binding in the brain of depressed patients
Authors: Kano Michiko , Fukudo Shin , Tashiro Atshushi , Utsumi Atshushi , Tamura Daisaku , Itoh Masatoshi , Iwata Ren , Tashiro Manabu , Mochizuki Hideki , Funaki Yoshihito , Kato Motohisa , Hongo Michio , Yanai Kazuhiko
Published: July 2004

The central histaminergic neuron system modulates the wakefulness, sleep–awake cycle, appetite control, learning and memory, and emotion. Previous studies have reported changes in neuronal histamine release and its metabolism under stress conditions in the mammalian brain. In this study, we examined, using positron emission tomography (PET) and [11C]‐doxepin, whether the histaminergic neuron system is involved in human depression. Cerebral histamine H1 receptor (H1R) binding was measured in 10 patients with major depression and in 10 normal age‐matched subjects using PET and [11C]‐doxepin. Data were calculated by a graphical analysis on voxel‐by‐voxel and ROI (region of interests) basis. Binding potential (BP) values for [11C]‐doxepin binding in the frontal and prefrontal cortices, and cingulate gyrus were significantly lower in the depressed patients than those in the normal control subjects. There was no area of the brain where [11C]‐doxepin binding was significantly higher in the depressed patients than in the controls. ROI‐based analysis also revealed that BP values for [11C]‐doxepin binding in the frontal cortex and cingulate gyrus decreased in proportion to self‐rating depressive scales scores. The results of this study demonstrate that depressed patients have decreased brain H1R binding and that this decrease correlates with the severity of depression symptoms. It is therefore suggested that the histaminergic neuron system plays an important role in the pathophysiology of depression and that its modulation may prove to be useful in the treatment of depression.
Is invasion success explained by the enemy release hypothesis?
Authors: Colautti Robert I. , Ricciardi Anthony , Grigorovich Igor A. , MacIsaac Hugh J.
Published: July 2004

A recent trend in invasion ecology relates the success of non‐indigenous species (NIS) to reduced control by enemies such as pathogens, parasites and predators (i.e. the enemy release hypothesis, ERH). Despite the demonstrated importance of enemies to host population dynamics, studies of the ERH are split – biogeographical analyses primarily show a reduction in the diversity of enemies in the introduced range compared with the native range, while community studies imply that NIS are no less affected by enemies than native species in the invaded community. A broad review of the invasion literature implies at least eight non‐exclusive explanations for this enigma. In addition, we argue that the ERH has often been accepted uncritically wherever (i) NIS often appear larger, more fecund, or somehow ‘better’ than either congeners in the introduced region, or conspecifics in the native range; and (ii) known enemies are conspicuously absent from the introduced range. However, all NIS, regardless of their abundance or impact, will lose natural enemies at a biogeographical scale. Given the complexity of processes that underlie biological invasions, we argue against a simple relationship between enemy ‘release’ and the vigour, abundance or impact of NIS.
Adaptation in an insect host–plant pathogen interaction
Authors: Cory Jenny S. , Myers Judith H.
Published: July 2004

Selection on parasites to adapt to local host populations may be direct or through other components of the system such as vectors or the food plant on which the parasite is ingested. To test for local adaptation of nucleopolyhedrovirus among island populations of western tent caterpillars, Malacosoma californicum pluviale, we compared virus isolates from three geographically distinct sites with different dominant host plants. Pathogenicity, speed of kill and virus production of each isolate were examined on the three food plants. Virus isolates from the two permanent host populations had the fastest speed of kill on the host plant from which they were isolated. This was not the case for a caterpillar population that goes extinct when populations are regionally low. Virus isolates on some plant species combined rapid speed of kill with high virus yield. Infection of hosts by mixed microparasite populations could facilitate local adaptation in response to differing food plant chemistry.
The behavioural ecology of personality: consistent individual differences from an adaptive perspective
Authors: Dall Sasha R. X. , Houston Alasdair I. , McNamara John M.
Published: July 2004

Individual humans, and members of diverse other species, show consistent differences in aggressiveness, shyness, sociability and activity. Such intraspecific differences in behaviour have been widely assumed to be non‐adaptive variation surrounding (possibly) adaptive population‐average behaviour. Nevertheless, in keeping with recent calls to apply Darwinian reasoning to ever‐finer scales of biological variation, we sketch the fundamentals of an adaptive theory of consistent individual differences in behaviour. Our thesis is based on the notion that such ‘personality differences’ can be selected for if fitness payoffs are dependent on both the frequencies with which competing strategies are played and an individual's behavioural history. To this end, we review existing models that illustrate this and propose a game theoretic approach to analyzing personality differences that is both dynamic and state‐dependent. Our motivation is to provide insights into the evolution and maintenance of an apparently common animal trait: personality, which has far reaching ecological and evolutionary implications.
Using the angle frequency method to detect signals of competition and predation in experimental time series
Authors: Sandvik Gunnar , Jessup Christine M. , Seip Knut L. , Bohannan Brendan J. M.
Published: July 2004

Identifying interactions among organisms is central to the study of ecology. The Angle Frequency Method (AFM) allows the detection of interactions in time series data. The AFM takes pairwise data plotted in phase diagrams and identifies signals (vector directions in phase diagrams) associated with particular interactions. Using microbial experimental systems consisting of predators (bacteriophage T4) and prey/competitors (strains of Escherichia coli), we demonstrate that the AFM can identify predator–prey and competitive interactions. The level of control afforded by such microbial experimental systems allows direct tests of the utility and robustness of the AFM. Signals of predation were distinct from signals of competition, with the strongest signal of predation corresponding to the collapse of the predator population at low prey densities. Signals of competition reflected the difference in competitive strength between the superior and the inferior competitors. In addition, the effects of invasion and resource enrichment on interactions in the laboratory communities were detectable using the AFM. Our analyses support results from model simulations and analyses of lake time series by identifying similar sets of signals characteristic of predation and competition, and demonstrate that the AFM is an effective tool in rigorous studies of time series.
Markov chain models predict the consequences of experimental extinctions
Authors: Timothy Wootton J.
Published: July 2004

Predicting the effects of species extinction is challenging because of ecosystem complexity. Using novel long‐term experimental deletions of two mussel species, I found that a multi‐species Markov chain model, which was parameterized under non‐experimental conditions, accurately predicted the effects of local extinction in an intertidal community. Systematically deleting species from the model predicted declining ecosystem biomass with reduced biodiversity, but relatively constant ecosystem resilience following disturbance. The effects of reduced biodiversity on these variables ranged widely, illustrating the need for species‐specific predictions of extinction effects. The results show that Markov chain models can be useful tools for predicting consequences of local species extinction without extensive experimentation.
Connectivity, scale‐dependence, and the productivity–diversity relationship
Authors: Chase Jonathan M. , Ryberg Wade A.
Published: July 2004

We surveyed freshwater ponds (localities) nested within watersheds (regions) to evaluate the relationship between productivity and animal species richness at different spatial scales. In watersheds where the ponds were relatively distant from one another (likely reducing the level of interpond dispersal of many organisms), we found a scale‐dependent productivity–diversity relationship; at local scales (among ponds), diversity was a hump‐shaped function of productivity, whereas at regional scales (among watersheds), diversity monotonically increased with productivity. Furthermore, this relationship emerged because there was a strong relationship between productivity and pond‐to‐pond species compositional differences. Alternatively, in watersheds where ponds were relatively close together (likely leading to higher rates of dispersal of many organisms), we found no scale‐dependence; diversity was a hump‐shaped function of productivity at both local and regional scales. Here, the relationship between species compositional dissimilarity and productivity was much weaker. We conclude that whether or not scale‐dependence is observed in productivity–diversity relationships will depend, at least in part, on the degree of connectivity among localities within regions.