bluuebunny/crossref_metadata_2025 huggingface dataset:
Pathology of bacterial gill disease: sequential development of lesions during natural outbreaks of disease
Authors: SPEARE D. J. , FERGUSON H. W. , BEAMISH F. W. M. , YAGER J. A. , YAMASHIRO S.
Published: April 2006

Abstract. This study examines the sequential pattern of bacterial colonization and the sequential development of branchial lesions during typical outbreaks of bacterial gill disease (BGD) among four groups of commercially reared rainbow trout fingerlings, Oncorhynchus mykiss (Richardson). During a 5‐month monitoring regime, prior to the onset of natural outbreaks of BGD, gill morphology of examined fish remained unaltered. Bacterial colonization of the gills was immediately preceded by several gill changes which were widely present in fish from all groups under study. These changes, which were detectable at the ultrastructural level only, included cytoplasmic blistering and degeneration of the microridges of the superficial filament epithelium, in addition to slight topographical irregularity of the filament tips suggestive of mild hyperplasia. Bacterial colonization began at these altered filament tips before spreading to more proximal regions of the filament and adjacent lamellar surfaces. Explosive increases in proportional morbidity and mortality coincided with the development of the following gill lesions: extensive bacterial colonization of lamellar surfaces, lamellar epithelial hydropic degeneration and necrosis, and lamellar oedema. In contrast, gill lesions such as lamellar fusion, epithelial hyperplasia and various metaplastic responses were detected later either as subacute (3–5 days) or chronic (7–14 days) changes. The role of epithelial necrosis as an important event during BGD is discussed as a possible mechanism leading to the development of sub‐acute and chronic gill lesions.
Instability of the major soluble antigen produced by Renibacterium salmoninarum
Authors: GRIFFITHS S. G. , LYNCH W. H.
Published: April 2006

Abstract. Using Western blot to examine the nature of soluble antigens produced by Renibacterium salmoninarum, it was found that the major 57‐kilodalton (kDa) antigen was unstable, SDS‐PAGE of extracellular product (ECP) fractions showed that degradation of the 57‐kDa protein increased with time and increased temperature. Several lower molecular mass pcptides accumulated temporarily from this degradation. Phenylmethylsulphonyl fluoride prevented breakdown of the 57‐kDa protein suggesting a scrine protease present in the ECPs was responsible. The results indicated that most, if not all, immunoreactive bands in ECP fractions, other than the 57‐kDa protein, arose as a result of degradation of this protein. Western blot analysis of two dimensional gels revealed that the presumptive proteolytic activity was associated with the 57‐kDa antigen and several of the apparent degradation products. Many common peptide fragments appeared to be generated from heat‐induced proteolysis of these protein moieties, confirming the familial relationship between much of the immunoreactive material in ECP fractions. The results suggested that the 57‐kDa antigen is autolytic. Western blot analysis of tissue samples from Atlantic salmon, Satmo solar L., infected with R. salmoninarum suggested that this lability of the 57‐kDa antigen also occurred in situ.
Macrophages and giant cells associated with a microsporidian parasite causing liquefaction of the skeletal muscle of the Norway pout, Trisopterus esmarkii (Nilsson)
Authors: PULSFORD A. , MATTHEWS R. A.
Published: April 2006

Abstract. A mature microsporidian infection of the skeletal muscles of Norway pout, Trisopterus esmarkii Nilsson, is described. Infected fish had noticeable liquefaction of the muscle. The white foci of infection contained packed microsporidian spores, and the surrounding infected muscle fibres were degenerate. The host response involved invasion by phagocytic cells and encapsulation by fibroblasts, forming large granulomas. Macrophages, 17–20 μm in diameter, were actively ingesting the spores and typically had a foamy cytoplasm. Some had formed Langhans‐type giant cells, 35 μm in diameter, with peripherally arranged nuclei. Sporulation was complete and appeared to have occurred within sporo‐ phorous vacuoles in which numerous spores were formed. Individual spores measured 2·8 × 1·5 μm and were characterized by irregular electron dense areas, a short polar filament with four coils and a large posterior vacuole with three inclusions.
Comparison of the effectiveness of three different growth media for primary isolation of Renibacterium salmoninarum from Atlantic salmon, Salmo salar L., broodfish
Authors: GUDMUNDSDóTTIR S. , HELGASON S. , BENEDIKTSDOTTIR E.
Published: April 2006

Abstract. The effectiveness of three different growth media KDM‐2, S‐KDM and S‐KDM‐C for primary isolation of Renibacterium salmoninarum was examined over a 14‐week incubation period. Kidney samples were taken from Atlantic salmon, Salmo salar L., broodfish returning to a sea ranch after 2 years at sea. Homogenized samples were inoculated onto two selective media, S‐KDM supplemented with lamb serum and S‐KDM‐C supplemented with activated charcoal. The third medium was the non‐selective serum supplemented KDM‐2. Renibacterium salmoninarum was isolated from 112 samples on one or more of the media used. Of all positive samples, 91% were positive on S‐KDM, 60% on S‐KDM‐C and 35% on KDM‐2. These results demonstrate that selectivity significantly enhances the isolation capacity of the medium and that supplementing with serum is significantly more effective than supplementing with charcoal.
Incubation time for the cultivation of Renibacterium salmoninarum from Atlantic salmon, Salmo salar L., broodfish
Authors: BENEDIKTSDÓTTIR E. , HELGASON S. , GUDMUNDSDÓTTIR S.
Published: April 2006

Abstract. Cultivation plates for the primary isolation of Renibacterium salmoninarum were incubated for 12 or more weeks over 3.5 years. The incubation time for sample of two groups of Atlantic salmon, Salmo solar L., broodfish on a selective agar medium is reported. Samples turned positive over a long time‐span. but the longest incubation time recorded for growth of the bacterium was 19 week. In one group, 79% of all positive samples form fish with macroscopic signs of bacterial kidney disease were positive after 6 weeks of incubation and the highest incidence of positive samples occured during the sixth week. From covertly infected fish within the same group.42% of sample were positive after 6 weeks of incubation, and the highest incidence of positive sample occured during the sixth and the ninth weeks of incubation. The growth pattern of the bacterium at primary isolation was characterized by a prolonged lag phase followed by a relatively growth in one week from microscopic to macroscopic size of the colonies.
The effect of glycyrrhizin on rainbow trout, Oncorhynchus mykiss (Walbaum), leucocyte responses
Authors: JANG S. I. , MARSDEN M. J. , KIM Y. G. , CHOI M. S. , SECOMBES C. J.
Published: April 2006

Abstract. Treatment of rainbow trout macrophages with glycyrrhizin (GL), an aqueous extract of licorice, Glycyrrhiza glabra, enhanced their respiratory burst activity. Maximal effects were seen using concentrations of 10–100 μml−1. GL also modulated trout lymphocytes, increasing proliferation responses to the mitogen phytohaemagglutinin some two‐fold over a range of GL concentrations. In addition, GL elicited the release of a macrophage activating factor (MAF) from head kidney leucocytes, as assessed by the ability of generated supernatants to increase respiratory burst activity of target macrophages. MAF activity was most apparent using 100 μg ml−1 GL to induce MAF release and a 48‐h incubation period with the target macrophages. Finally, GL was shown to enhance the release of MAF in response to the mitogen concanavalin A. The possible use of GL as a stimulant of fish innate defences is discussed.
Attitudes toward participation in a pregnancy and child cohort study
Authors: Daniels Julie L. , Savitz David A. , Bradley Chyrise , Dole Nancy , Evenson Kelly R. , Eucker Barbara , Herring Amy H. , Siega‐Riz Anna Maria , Thorp John M.
Published: April 2006

While epidemiological studies aim for high participation rates, it is becoming increasingly difficult to recruit and retain participants in lengthy observational studies. We surveyed women who recently participated in the Pregnancy, Infection, and Nutrition Study during their pregnancy to learn more about what initially motivated them to participate in the study, their attitudes about the study protocol, and whether they would allow their child to participate in future studies. Most women were motivated by their interest in science and learning about their pregnancy. In general, women felt quite comfortable with most aspects of the study. Consent forms, telephone interviews and self‐administered questionnaires were the most acceptable components of the study, but even specimen collection was well tolerated by this cohort. Women were less comfortable with the possibility of their child participating in future research. This survey confirmed that once women are enrolled, they tend to be willing to complete most components of an intensive study, suggesting that initial efforts for recruitment are most important.
Arginase activity in a murine macrophage cell line (RAW264.7) stimulated with lipopolysaccharide from Actinobacillus actinomycetemcomitans
Authors: Sosroseno W. , Musa M. , Ravichandran M. , Fikri Ibrahim M. , Bird P. S. , Seymour G. J.
Published: April 2006

Aims:  The aim of the present study was to determine whether or not lipopolysaccharide from Actinobacillus actinomycetemcomitans could stimulate arginase activity in a murine macrophage cell line (RAW264.7 cells).

Methods:  RAW264.7 cells were treated with A. actinomycetemcomitans‐lipopolysaccharide or lipopolysaccharide from Escherichia coli for 24 h. The effect of polymyxin B, l‐norvaline, dl‐norvaline, dexamethasone and cytokines (interferon‐γ and interleukin‐4) on arginase activity in A. actinomycetemcomitans‐lipopolysaccharide‐stimulated cells was also determined. The cells were pretreated with anti‐CD14, anti ‐toll‐like receptor 2, or anti‐toll‐like receptor 4 antibody prior to stimulation with A. actinomycetemcomitans‐lipopolysaccharide. Arginase activity was determined by a colorimetric assay.

Results:  A. actinomycetemcomitans‐lipopolysaccharide stimulated arginase activity in RAW264.7 cells in a dose‐dependent manner, but was less potent than E. coli‐lipopolysaccharide. Polymyxin B and l‐norvaline, but not dl‐norvaline, blocked the arginase activity in A. actinomycetemcomitans‐lipopolysaccharide‐stimulated cells. Dexamethasone and interleukin‐4 but not interferon‐γ augmented arginase activity in A. actinomycetemcomitans‐lipopolysaccharide‐stimulated cells. Treatment of the cells with anti‐CD14 and anti‐toll‐like receptor 4 but not anti‐toll‐like receptor 2 antibody decreased arginase activity in A. actinomycetemcomitans‐lipopolysaccharide‐stimulated cells.

Conclusion:  The results of the present study suggest that lipopolysaccharide from A. actinomycetemcomitans via CD14/toll‐like receptor 4 complex molecules and the regulatory control of glucocorticoid and cytokines may stimulate arginase activity in RAW264.7 cells.
Inhibitory effects of macrocarpals on the biological activity of Porphyromonas gingivalis and other periodontopathic bacteria
Authors: Nagata H. , Inagaki Y. , Yamamoto Y. , Maeda K. , Kataoka K. , Osawa K. , Shizukuishi S.
Published: April 2006

Background/aims:  Macrocarpals, which are phloroglucinol derivatives contained in eucalyptus leaves, exhibit antimicrobial activity against a variety of bacteria including oral bacteria. This study examined effects of macrocarpals A, B, and C on periodontopathic bacteria, especially Porphyromonas gingivalis.

Methods:  Macrocarpals A, B, and C were purified from a 60% ethanol‐extract of Eucalyptus globules leaves. To investigate antibacterial activity, representative periodontopathic bacteria were cultured in media with or without various amounts of macrocarpals; subsequently, the optical density at 660 nm was measured. Macrocarpal inhibition of P. gingivalis Arg‐ and Lys‐specific proteinases was assessed by spectrofluorophotometric assay and sodium dodecyl sulfate‐polyacrylamide gel electrophoresis analysis. The effect of macrocarpals on P. gingivalis binding to saliva‐coated hydroxyapatite beads was examined with 3H‐labeled P. gingivalis.

Results:  Growth of P. gingivalis was inhibited more strongly than growth of Prevotella intermedia or Prevotella nigrescens and Treponema denticola by macrocarpals, however, Actinobacillus actinomycetemcomitans and Fusobacterium nucleatum were much more resistant. Macrocarpals inhibited P. gingivalis Arg‐ and Lys‐specific proteinases in a dose‐dependent manner. The enzyme‐inhibitory effect of macrocarpals was confirmed by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis analysis in which hemoglobin degradation by P. gingivalis proteinase was inhibited by macrocarpals. P. gingivalis binding to saliva‐coated hydroxyapatite beads was also strongly attenuated by macrocarpals.

Conclusions:  Macrocarpals A, B and C demonstrated antibacterial activity against periodontopathic bacteria. Among tested bacteria, P. gingivalis displayed the greatest sensitivity to macrocarpals; additionally, its trypsin‐like proteinase activity and binding to saliva‐coated hydroxyapatite beads were inhibited by macrocarpals. These results indicate that eucalyptus leaf extracts may be useful as a potent preventative of periodontal disease.
Antimicrobial susceptibility and molecular analysis of Enterococcus faecalis originating from endodontic infections in Finland and Lithuania
Authors: Reynaud af Geijersstam A. H. , Ellington M. J. , Warner M. , Woodford N. , Haapasalo M.
Published: April 2006

Background/aims:  Enterococcus faecalis strains with multiple antibiotic resistances can cause infections that are difficult to treat. The microbial flora in treatment‐resistant apical periodontitis is dominated by E. faecalis, and is a potential source of infections at other sites.

Material and methods:  Sensitivities to a range of antibiotics were determined for 59 endodontic E. faecalis isolates from Finland and Lithuania. The DNA sequence of the gene responsible for the species' intrinsic quinupristin‐dalfopristin resistance, lsa, was determined from two isolates with diminished resistance. Four pairs of isolates from the same root canal were typed by pulsed‐field gel electrophoresis.

Results:  A high prevalence of resistance to rifampicin was found, whereas all isolates were susceptible or showed intermediate susceptibility to penicillin and ampicillin and four isolates were unusually susceptible to cefotaxime. No vancomycin or high‐level gentamicin resistance was detected. Nine of 59 isolates were susceptible to quinupristin‐dalfopristin. A fully quinupristin‐dalfopristin‐susceptible isolate also susceptible to clindamycin produced a truncated Lsa polypeptide, and an isolate with borderline quinupristin‐dalfopristin‐susceptibility had mutations proximal to the predicted ribosomal binding site. Pulsed‐field gel electrophoresis showed that the same root canal could harbor two different strains of E. faecalis during the course of the same infection.

Conclusion:  Despite the differing antibiotic usage in Finland and Lithuania, E. faecalis from endodontic infections in these countries showed similar susceptibility patterns with levels of resistance considered typical for the species, and decreased resistance to clindamycin and quinupristin‐dalfopristin as well as lesions in the lsa gene which were similar to those described in other clinical isolates.
Human T‐cell responses to oral streptococci in human PBMC‐NOD/SCID mice
Authors: Salam M. A. , Nakao R. , Yonezawa H. , Watanabe H , Senpuku H.
Published: April 2006

We investigated cellular and humoral immune responses to oral biofilm bacteria, including Streptococcus mutans, Streptococcus anginosus, Streptococcus sobrinus, and Streptococcus sanguinis, in NOD/SCID mice immunized with human peripheral blood mononuclear cells (hu‐PBMC‐NOD/SCID mice) to explore the pathogenicity of each of those organisms in dental and oral inflammatory diseases. hu‐PBMC‐NOD/SCID mice were immunized by intraperitoneal injections with the whole cells of the streptococci once a week for 3 weeks. FACS analyses were used to determine the percentages of various hu‐T cell types, as well as intracellular cytokine production of interleukin‐4 and interferon‐γ. Serum IgG and IgM antibody levels in response to the streptococci were also determined by enzyme‐linked immunosorbent assay. S. anginosus induced a significant amount of the proinflammatory cytokine interferon‐γ in CD4+ and CD8+ T cells in comparison with the other streptococci. However, there was no significant differences between the streptococci in interleukin‐4 production by CD4+ and CD8+ T cells after inoculation. Further, S. mutans significantly induced human anti‐S. mutans IgG, IgG1, IgG2, and IgM antibodies in comparison with the other organisms. In conclusion, S. anginosus up‐regulated Th1 and Tc1 cells, and S. mutans led to increasing levels of their antibodies, which was associated with the induction of Th2 cells. These results may contribute to a better understanding of human lymphocyte interactions to biofilm bacteria, along with their impact on dental and mucosal inflammatory diseases, as well as endocarditis.
In vitro antifungal susceptibility to six antifungal agents of 229 Candida isolates from patients with diabetes mellitus
Authors: Manfredi M. , McCullough M. J. , Polonelli L. , Conti S. , Al‐Karaawi Z. M. , Vescovi P. , Porter S. R.
Published: April 2006

The most common antifungal drugs in current clinical use for the treatment of oral candidosis are polyenes and azoles, mainly used topically. Poor glycaemic control in association with other local factors, such as the presence of oral dental prostheses, salivary pH, salivary flow rate and tobacco habits, may lead to the development of oral candidosis. Topical antifungal agents are frequently used to prevent the development of candidal infections in patients with poor metabolic control, particularly in the elderly wearing dentures. The aim of this study was to assess the antifungal susceptibility of Candida isolates to six antifungal agents using a commercially available kit, Fungitest®. The isolated were collected from patients affected by diabetes mellitus from two different geographic localities (London, UK, and Parma, Italy) and from a group of healthy non‐diabetic subjects. No differences in antifungal susceptibility to the six agents tested were observed between Candida isolates from diabetic and non‐diabetic subjects. However, differences were observed between the two geographically different diabetes mellitus populations. Oral yeast isolates from diabetes mellitus patients in the UK more often displayed resistance or intermediate resistance to fluconazole (P = 0.02), miconazole (P < 0.0001), and ketoconazole (P = 0.01) than did isolates from diabetes mellitus patients in Italy. In addition, more C. albicans isolates were found in diabetic and non‐diabetic subjects that were susceptible to fluconazole (P = 0.0008 and P = 0.01, respectively) than non‐albicans isolates. The difference in the antifungal resistance of isolates from the two populations of diabetes mellitus patients may be related to differences in the therapeutic management of candidal infections between the two centres.
Response of human macrophage‐like cells to stimulation by Fusobacterium nucleatum ssp. nucleatum lipopolysaccharide
Authors: Grenier D. , Grignon L.
Published: April 2006

Monocytes/macrophages are key members of the innate immune system and are present in higher numbers in active periodontal lesions than in inactive sites. The aim of this study was to characterize the response of human monocyte U937 cells, differentiated into adherent macrophages by treatment with phorbol‐12‐myristate 13‐acetate, to stimulation by Fusobacterium nucleatum ssp. nucleatum lipopolysaccharide. Attachment of 3H‐lipopolysaccharide to macrophage‐like cells was partially inhibited by anti‐CD14 and anti‐TLR4 polyclonal antibodies. Fusobacterial lipopolysaccharide did not cause cell apoptosis or block apoptosis induced by camptothecin. Lipopolysaccharide up‐regulated the secretion of the pro‐inflammatory cytokines interleukin‐1β, interleukin‐6, and tumor necrosis factor‐α as well as the chemokine interleukin‐8 by macrophage‐like cells. In addition, it increased phospholipase C and D activities, which likely contributed to the high levels of prostaglandin E2 detected in the cell culture supernatant. Lastly, the amount of matrix metalloproteinase‐9 produced by macrophage‐like cells was significantly increased by the lipopolysaccharide treatment. Interestingly, fusobacterial cells acquired matrix metalloproteinase‐9 activity following incubation in the presence of the culture supernatant of lipopolysaccharide‐stimulated macrophage‐like cells. In summary, the lipopolysaccharide of F. nucleatum ssp. nucleatum has a large array of biological effects on macrophage‐like cells. This monocytic responsiveness to lipopolysaccharide may be a key regulator of periodontitis.
Phenotypic and genotypic identification of enterococci isolated from canals of root‐filled teeth with periapical lesions
Authors: Pinheiro E. T. , Anderson M. J. , Gomes B. P. F. A. , Drucker D. B.
Published: April 2006

The objectives of the present study were to identify enterococcal species isolated from the canals of root‐filled teeth with periapical lesions using biochemical and molecular techniques, and to investigate the genetic diversity of the isolates. Twenty‐two Enterococcus strains, isolated from the canals of root‐filled teeth with persisting periapical lesions, were identified to species level using rapid ID 32 STREP galleries and partial 16S rDNA sequencing. To subtype the strains, genomic DNA from the isolates was analyzed by pulsed field gel electrophoresis (PFGE) after digestion with SmaI. Intragenic regions of two genes, ace and salA, were sequenced for further differentiation of the isolates. All strains were identified as Enterococcus faecalis by both commercial kit and partial 16S rDNA sequencing. PFGE with SmaI of 22 isolates demonstrated 18 macrorestriction profiles, whereas 13 distinct genotypes were identified after analysis of the ace and salA composite sequences. Most of the isolates from distinct patients had different PFGE profiles. Moreover, in two cases, different E. faecalis strains were found in different root‐filled teeth from the same mouth. E. faecalis was the only enterococcal species isolated from the canals of root‐filled teeth with periapical lesions. Genetic heterogeneity was observed among the E. faecalis isolates following PFGE and sequence‐based typing method. Furthermore, the genetic diversity within root canal strains was similar to previous reports regarding E. faecalis isolates from different clinical and geographic origins.
IgG monitoring to identify the risk for development of infection in heart transplant recipients
Authors: Sarmiento E. , Rodriguez‐Molina J.J. , Fernandez‐Yañez J. , Palomo J. , Urrea R. , Muñoz P. , Bouza E. , Fernandez‐Cruz E. , Carbone J.
Published: April 2006

Abstract: Infectious complication represents a significant source of morbidity and mortality in heart transplant recipients. To assess humoral immunity markers that can predict the development of infection, 38 consecutive recipients of heart transplants performed at a single center were prospectively studied. Induction therapy included daclizumab. Immunoglobulin (IgG, IgA, IgM) and complement factors (C3, C4, and factor B) were performed by nephelometry in peripheral blood samples obtained before transplantation, and 7 days and 1 month after transplantation. During a mean follow‐up of 16.9 months, 13 patients had at least one episode of infection (34.2%). Eight of these were cytomegalovirus (CMV) infections treated with intravenous ganciclovir, 2 were bacterial pneumonia, 1 patient had bacterial septicemia, 1 patient had urinary tract infection, and 1 patient had pulmonary nocardiosis. No significant association was found between infection and age, sex, immunosuppression, CMV serostatus of donor and recipient, or treated rejection episodes. Pre‐transplant IgG (below median value=1140 mg/dL; relative risk [RR] 3.69; 95% confidence interval [CI] 1.01–13.54; P=0.04) and post‐transplant IgG levels at day 7 (below median value=679 mg/dL; RR 11.21; CI 1.04–89.48; P=0.022) were associated with an increase in the risk for developing infections. Early monitoring of immunoglobulin levels might help to identify the risk for developing infection in heart transplantation.
Actions of Somatostatin on Rat Neocortical Neurons in vitro
Authors: Vidal C. , Zieglgänsberger W.
Published: April 2006

The effects of somatostatin (SS14) on neocortical neurons of the rat were investigated in an in vitro slice preparation. Intracellular recordings were performed in neurons (n=30) in layers 2 and 3 of the frontal cortex.

Iontophoretically applied SS14 reduced the responses evoked by iontophoretically applied L‐glutamate (GLU) and γ‐aminobutyric acid (GABA). The blocking effect of SS14 was apparent 1–2 min after onset of SS14 application and recovery required 2–3 min. The conductance increase evoked by GLU or GABA was reduced by SS14. In the majority of neurons, SS14 did not produce any measurable changes in passive membrane properties, spike threshold or on orthodromically evoked synaptic potentials. In 5 cells, SS14 induced a slight hyperpolarization (<3 mV).

These results lend further support to claims that SS14 plays a neuromodulatory role in the neocortex.
Modular Connections between Areas V2 and V4 of Macaque Monkey Visual Cortex
Authors: Zeki S. , Shipp S.
Published: April 2006

We have studied the connections between two visual areas of macaque monkey cortex, V2 and V4, by injecting wheat‐germ agglutinin horseradish peroxidase (HRP‐WGA) into V4 and examining the distribution of labelled cells and terminals in V2, in relation to its characteristically striped cytochrome oxidase architecture. The cells projecting from V2 to V4 are arranged in bands and the number of bands per cycle of cytochrome oxidase stripes varies (one cycle consists of a thin stripe, a thick stripe and two interstripes). In the Type 1 connectivity pattern, there is just one band per cycle, centred over the thin stripes but normally spreading into the neighbouring interstripes. In the Type 2 connectivity pattern there are two bands per cycle, generally rather narrower and centred over the interstripes. Thick stripes are mostly free of labelled cells. The return projection from V4 to V2, whilst being concentrated in the vicinity of the labelled cells, is more diffusely distributed and invades the territory of all the stripes.
The Influence of Magnesium lons on the NMDA Mediated Responses of Ventral Rhythmic Neurons in the Spinal Cord of Xenopus Embryos
Authors: Soffe S. R. , Roberts Alan
Published: April 2006

Xenopus embryos immobilized in tubocurarine respond to natural skin stimulation with fictive swimming. This can also occur in saline without Mg2+ and is blocked by NMDA antagonists. Ventral spinal cord neurons which are rhythmically active during swimming are depolarized by bath applied N‐methyl‐D‐aspartate (NMDA) (in 1 μM tetrodotoxin (TTX) to block indirect effects). By using current clamp techniques this depolarization is shown to be partially blocked by 0.5 and 1 mM Mg2+ in a voltage‐dependent manner similar to that described in cultured neurons.

Mg2+ partially and reversibly reduces the slow NMDA‐mediated component of excitatory post‐synaptic potentials (EPSPs) in ventral neurons. However, in 1 mM Mg2+ fictive swimming can still be evoked by natural stimulation. The frequency of swimming is slightly lower than in nominally 0 mM Mg2+, but the pattern of ventral root activity and synaptic drive to ventral neurons seems little affected. Fictive swimming can also be induced by applying NMDA to spinal preparations. In 0 mM Mg2+, such rhythmic activity is unstable and transient over a narrow NMDA concentration range. In 0.5 mM Mg2+, continuous rhythmic activity is induced over a wide range of NMDA concentrations. Lower spinal preparations need higher NMDA concentrations to induce activity.

We conclude that the neurons rhythmically active in swimming have NMDA receptor channels which show a voltage dependent block in the presence of Mg2+. However, while Mg2+ exerts a powerful stabilizing influence on rhythmic activity induced in spinal embryos by exogenous NMDA, its influence on ‘naturally’ evoked fictive swimming is less clear. The fictive swimming machinery in the brain and spinal cord can produce stable swimming with or without Mg2+ induced voltage dependency of the NMDA channels.
Localization and Quantitative Autoradiography of Glutamatergic Ligand Binding Sites in Chick Brain
Authors: Henley Jeremy M. , Moratallo Rosario , Hunt Stephen P. , Barnard Eric A.
Published: April 2006

The anatomical localization of glutamate receptor subtype‐selective ligand binding sites was investigated in 1‐day‐old chick brain using quantitative autoradiography. Under the conditions used, the regional distributions of [3H]glutamate, [3H]AMPA (a selective quisqualate receptor ligand) and [3H]kainate binding sites are manifestly different. [3H]L‐glutamate binding is densely localized in the telencephalon, particularly in the neostriatum (2.8 pmol/mg protein). In addition, [3H]L‐glutamate labels the thalamus, the nucleus mesencephalicus lateralis pars dorsalis, the superficial layers of the optic tectum and the molecular layer of the cerebellum. [3H]AMPA binding sites are most densely localized in the hippocampus (0.90 pmol/mg protein), with an otherwise relatively uniform distribution of binding within the telencephalon. [3H]AMPA also labels the striatum griseum et fibrosum superficiale of the optic tectum and the molecular layer of the cerebellum. [3H]Kainate binding sites are extremely densely packed in the molecular layer of the cerebellum (10 pmol/mg protein). Other regions of [3H]kainate binding include the hyperstriatum and the thalamus. The binding of the NMDA receptor channel blocker [3H]MK‐801 is increased in the presence of 1 mM L‐glutamate. [3H]MK‐801 binding is generally widespread in the telencephalon but is notably absent from the ectostriatum. No evidence of [3H]MK‐801 binding sites was detected in the cerebellum, even in the presence of 1 mM L‐glutamate. The relatively high densities and the well‐defined localizations of the glutamate receptor subtype binding sites suggest that chick brain provides a useful system for the further study of excitatory amino acid receptors.
Omissions of Auditory Stimuli May Activate Frontal Cortex
Authors: Joutsiniemi S.‐L. , Hari R.
Published: April 2006

We measured, with a 7 channel SQUID gradiometer, cerebral magnetic responses, time‐locked to unpredictable and infrequent stimulus omissions (10% of stimuli) in an otherwise regular sequence of short tones, repeated once every 510 ms. In 10 out of 12 subjects broad responses with multiple peaks and with wide interindividual variability were detected. Attention clearly increased the observed signals. The magnetic field patterns evoked by stimulus omissions could be explained by single current dipoles, whose locations agree with activation of the posterolateral frontal cortex.
Roles of Glycinergic Inhibition and N‐Methyl‐D‐Aspartate Receptor Mediated Excitation in the Locomotor Rhythmicity of One Half of the Xenopus Embryo Central Nervous System
Authors: Soffe S. R.
Published: April 2006

Recent investigations into the neural basis for swimming in Xenopus embryos have pointed to central roles played by N‐methyl‐D‐aspartate (NMDA) receptor‐mediated excitation acting within and glycinergic reciprocal inhibition acting between motor systems for the muscular antagonists on the two sides of the CNS. A ‘reduced’ preparation consisting of only one half of the CNS divided sagittally along its midline is used here to examine the basis for rhythmicity within each side in the absence of reciprocal connections. Divided preparations transected rostrally at levels between the otic capsule and the obex can all generate a rhythmic pattern of motor discharge similar to that which underlies swimming. All rhythm generation is blocked by the NMDA antagonist (+/ ‐)‐2‐amino‐5‐phosphonovaleric acid (APS) at 20 μM. However, neither glycinergic nor GABAergic inhibition is required for a basic rhythmicity since some rhythm persists in the presence of 10 μM strychnine and 50 μM bicuculline, though it is no longer sustained. In the divided spinal cord alone, rhythm generation requires extracellular Mg2+. If the most caudal segment of the divided hindbrain is left attached, extracellular Mg2+ is required only if strychnine is present. If more of the hindbrain is included, extracellular Mg2+ is no longer necessary for rhythm generation even in the presence of strychnine.

It seems that rhythm generation by a single side of the spinal cord requires NMDA receptor‐mediated excitation together with the voltage dependency conferred on it by extracellular Mg2+, but not inhibition in order to occur. As more of the hindbrain is left attached, the requirement for extracellular Mg2+ becomes progressively less strong. For sustained rhythm generation, one side of the CMS requires both excitation and glycinergic inhibition.
Anatomical and Physiological Properties of GABAergic Neurotransmission in Organotypic Slice Cultures of Rat Hippocampus
Authors: Streit Peter , Thompson Scott M. , Gähwiler Beat H.
Published: April 2006

The anatomical and physiological properties of GABAergic inhibitory neurotransmission were investigated in organotypic slice cultures of rat hippocampus. Interneurons and terminal‐like elements containing GABA‐like immunoreactivity were numerous in tissue kept for 13–26 days in culture and showed a similar morphology and distribution to those known from investigations on the hippocampal formation in situ. Furthermore, after 8–30 days in culture, spontaneous and evoked IPSPs were observed in all CA3 pyramidal cells tested, resulting from an increase in chloride conductance, and were shown to be mediated by activation of GABA receptors. No functional decrement in the efficacy of GABAergic inhibitory synaptic transmission following chronic isolation and long‐term maintenance in vitro was noticed. In particular, neither the magnitude of the synaptic conductance underlying the inhibitory postsynaptic currents nor its reversal potential varied with time in culture. Taken together, the present physiological and immunohistochemical data show that GABAergic inhibition is well expressed in organotypic hippocampal slice cultures and is maintained over periods of at least 4 weeks in vitro.
Ultrastructural Characterization of Adrenocorticotrope Hormone (ACTH) Immunoreactive Fibres in the Mesencephalic Central Grey Substance of the Rat
Authors: Buma Pieter , Veening Jan , Nieuwenhuys Rudolf
Published: April 2006

The fine structural localization of fibres immunoreactive for the adrenocorticotrope hormone (ACTH) was studied in the mesencephalic central grey substance (MCG) of the male Wistar rat. Light microscopically, varicose ACTH‐immunoreactive fibres were found throughout the MCG in a dorsal, lateral and ventral, periventricular position. Electron microscopically, the immunoreactivity was most prominent in the direct vicinity of electron‐dense secretory granules in axonal varicosities, and, although to a lower degree, around other cytoplasmic organelles such as electron‐lucent synaptic vesicles, mitochondria and microtubules. With serial section analysis two types of ACTH‐immunoreactive varicosity were discerned. The first type is large, contains many, small electron‐lucent synaptic vesicles, that are located in the vicinity of a morphologically well‐defined synaptic contact. In this type of varicosity, large dense‐core secretory granules are scarce. Immunoreactivity is low or absent, particularly near the active zone. The second type is strongly immunoreactive. It always contains many large, dense‐core secretory granules; electron‐lucent vesicles are rare. The smaller varicosities of this type never make synaptic contacts, but a few of the larger varicosities have synaptic contacts with dendrites of MCG cells.
Absence of Wallerian Degeneration does not Hinder Regeneration in Peripheral Nerve
Authors: Lunn E. R. , Perry V. H. , Brown M. C. , Rosen H. , Gordon S.
Published: April 2006

Wallerian degeneration of the distal stump of a severed peripheral nerve involves invasion by myelomonocytic cells, whose presence is necessary for destruction of myelin and for initiating mitosis in Schwann cells (Beuche and Friede, 1984). Degeneration of the distal ends of the axons themselves is assumed to occur by autolytic mechanisms. We describe a strain of mice (C57BL/6/Ola) in which leucocyte invasion is slow and sparse. In these mice, confirming Beuche and Friede, myelin removal is extremely slow. A new finding is that axon degeneration is also very slow. This is a consequence of lack of recruitment of myelomonocytic cells for if such recruitment is prevented in other mouse strains by a monoclonal antibody against the complement type 3 receptor (Rosen and Gordon, 1987) axon degeneration is again slowed. We have also, surprisingly, found that nerve regeneration in the C57BL/6/Ola mice is not impeded by the presence of largely intact axons in the distal stump and absence of recruited cells, myelin debris and the absence of Schwann cell mitosis.
Degenerative Changes in Forebrain Cholinergic Nuclei Correlate with Cognitive Impairments in Aged Rats
Authors: Fischer Walter , Gage Fred H. , Björklund Anders
Published: April 2006

Degenerative changes in the forebrain cholinergic nuclei have been studied morphometrically in behaviourally characterized aged female Sprague‐Dawley rats. In all regions analysed (medial septum, diagonal band of Broca, nucleus basalis, and striatum) the acetylcholinesterase‐positive neurons were reduced in both size and number in the aged (24‐months‐old) rats as compared to the young (3‐months‐old) controls. The overall reduction in cell size amounted to between 20 and 30% and the overall reduction in cell number to between 27 and 45%. Impairment in learning and/or memory performance in the aged rats, as assessed in the Morris' water‐maze task, was significantly correlated with both cholinergic cell size and cell number in the medial septum, and with cholinergic cell number in the diagonal band of Broca and in the striatum. In the nucleus basalis there was a trend in the same direction but it did not reach significance. In contrast to these degenerative changes in the cell body regions, no significant differences in cortical or hippocampal choline acetyltransferase activity were detected biochemically between the young and the aged rats, and the enzyme activity levels did not correlate with the degree of behavioural impairment in the aged rats.

The present results provide evidence that all major forebrain cholinergic cell groups undergo degenerative changes with age in the rat, and that the most severe changes are found in those rats which display the most profound spatial learning impairments. Despite the severe changes at the cell body level, however, the choline acetyltransferase activity in the cortical projection areas are affected only to a minor degree, perhaps as a result of functional compensatory changes at the terminal level.
The Topographic Organization and Axis of Projection within the Visual Sector of the Rabbit's Thalamic Reticular Nucleus
Authors: Crabtree John W. , Killackey Herbert P.
Published: April 2006

The organization of the visual field representation within the thalamic reticular nucleus (TRN) of the rabbit was studied. Focal injections of horseradish peroxidase (HRP) and/or [3H]proline were made into visuocortical areas V1 and V2 and the dorsal lateral geniculate nucleus (dLGN). The resultant labelling in the thalamus was analysed.

A single injection in V1 or V2 results in a single zone of terminal label within the TRN that is restricted to the dorsocaudal part of the sheet‐like nucleus. In comparisons of the zones of label following injections at two different cortical sites in V1, a medial to lateral shift in label across the thickness of the TRN sheet is accompanied by a medial to lateral shift in label in the dLGN; a dorsal to ventral shift in label within the plane of the TRN sheet is accompanied by a dorsal to ventral shift in label in the dLGN. Thus, like the dLGN the TRN receives a precise topographic projection from V1. In reconstructions from horizontal sections the zones of label within the TRN resemble ‘slabs’, which lie within the plane of the nucleus parallel to its borders. Thus, the slabs of visuocortical terminals and reticular dendrites are similarly oriented. As revealed by the orientation of the slabs, the lines of projection representing points in visual space are represented by the oblique rostrocaudal dimension of the TRN.

Injections restricted to V1 produce terminal labelling that is confined to the outer two‐thirds of the TRN across its thickness, whilst those involving V2 result in terminal labelling within the inner one‐third of the nucleus. Thus, the adjacent cortical areas V1 and V2 project in a continuous fashion across the mediolateral dimension of the TRN. The organization of the map within the TRN, which was revealed by visuocortical injections, was confirmed by the pattern of retrograde labelling within the nucleus following geniculate injections of HRP.

On the basis of these findings and those in other mammalian species, two major conclusions can be reached that alter our view of the TRN. First, rather than mapping onto the whole nucleus in a continuous fashion, the cortical projection to the TRN has significant discontinuities. Second, rather than integrating efferents from widespread cortical areas, the reticular dendrites are related to focal areas of cortex.
Noradrenaline‐Induced Inhibition of Voltage‐Sensitive Calcium Currents in NG108‐15 Hybrid Cells
Authors: Docherty R. J. , McFadzean I.
Published: April 2006

The effect of noradrenaline (NA) on voltage‐sensitive calcium currents in neuroblastoma × glioma hybrid (NG108‐15) cells has been studied using a whole‐cell clamp technique. NA inhibited calcium current. The EC50 for NA induced inhibition was 177 nM. The NA receptor involved had a similar pharmacological profile to α2 adrenoreceptors but did not respond to clonidine. NA inhibited calcium current by an amount which depended on both membrane potential and current amplitude. When current‐dependent inactivation of calcium current was maximal, NA was without effect. The data are consistent with a mechanism where NA enhances current‐dependent inactivation.
A Novel Type of Glial Cell Associated with Nodes of Ranvier in Rat Optic Nerve
Authors: Miller Robert H. , Fulton Barbara P. , Raff Martin C.
Published: April 2006

Using Golgi impregnation and intracellular injection of horseradish peroxidase, we show that the adult rat optic nerve contains two distinct types of astrocyte‐like glial cells: one has mainly radially oriented processes that terminate on blood vessels or on the pial surface; the other has mainly longitudinally oriented processes that associate with, and often terminate at, nodes of Ranvier, but do not end on blood vessels or the pial surface. The sequence of appearance of the two types of glial cells in the developing nerve, taken together with previous immunocytochemical findings, suggests that these cells may correspond to the two types of astrocytes previously described in cultures of perinatal optic nerve cells—those with mainly radially oriented processes corresponding to type‐1 astrocytes and those with mainly longitudinally oriented processes corresponding to type‐2 astrocytes. To our knowledge, this is the first description of a class of central nervous system (CNS) glial cell whose processes are primarily associated with nodes of Ranvier.
Expansion of the Central Hindpaw Representation Following Fetal Forelimb Removal in the Rat
Authors: Killackey Herbert P. , Dawson Douglas R.
Published: April 2006

We provide evidence that prenatal removal of a rat forelimb results in both a disruption of the anatomical representation which would normally correspond to the forepaw and in an enlargement of the adjacent hindpaw representation in the brainstem and cortex. This enlargement, which in some cases is as much as 100%, only occurs following complete forelimb amputation on embryonic day 17 (E17) or earlier. This coincides with the age at which forepaw afferents first arrive in the brainstem, suggesting to us that the expansion is permitted in part because ingrowing hindpaw afferents are in the presence of cuneate cells which have never been previously innervated; in animals older than E17, the expansion is prohibited by either an intrinsic age‐dependent change in the cuneate cells, or a change imposed upon them by forelimb afferents.

The number of cells in dorsal root ganglia subserving the expanded hindpaw areas does not differ from normal suggesting that the expansion of hindpaw territory within the brainstem reflects an increased terminal arborization by a normal complement of primary hindpaw afferents.

We interpret the cortical enlargement to be an upstream reflection of the brainstem events. In cortex, the enlargement seems to result from an invasion of the dysgranular cortex by thalamic afferents arising from the ventral posterior nucleus.
Involvement of D1 and D2 Dopamine Receptors in the Control of Horizontal Cell Electrical Coupling in the Turtle Retina
Authors: Piccolino M. , Demontis G. , Witkovsky P. , Strettoi E. , Cappagli G. C. , Porceddu M. L. , De Montis M. G. , Pepitoni S. , Biggio G. , Meller E. , Bohmaker K.
Published: April 2006

We studied the actions of D1 and D2 dopamine agonists and antagonists on the coupling of horizontal cell axons in the turtle retina by a combinationn of pharmacological and electrophysiological methods. Both D1 and D2 receptors were identified in membrane fractions by radioligand binding using [3H]‐SCH 23390 and [3H]‐spiperone, respectively. The KD of both receptor classes were identical (0.21 nM) but D1 receptor density exceeded that of D2 receptors by more than four‐fold. D1 agonists increased the activity of adenylate cyclase in a dose‐dependent manner, whereas D2 agonists were without significant effect by themselves, nor did D2 antagonists block the D1‐mediated increase in adenylate cyclase activity. Intracellular recordings and Lucifer Yellow dye injections were used to characterize the modifications of the receptive field profile of horizontal cell axons (H1AT) exposed to different pharmacological agents. Dopamine or D1 agonists (0.05–10 μM) induced a marked constriction of the H1AT receptive field, whereas D2 agonists elicited a small expansion of the receptive field. However, in the presence of a D1 antagonist, as well as IBMX to inhibit phosphodiesterase, D2 agonists (10–70 μM) induced a marked increase in the receptive field profile. These results indicate that both D1 and D2 dopamine receptors play a role in shaping the receptive field profile of the horizontal cell axon terminal in the turtle retina.