High-affinity uptake of γ-aminobutyric acid in cultured glial and neuronal cells

Both glial and neuronal cells maintained in primary culture were found to accumulate [³H]GABA by an efficient high-affinity uptake system (apparentK _m=9 M,V _max=0.018 and 0.584 nmol/mg/min, respectively) which required sodium ions and was inhibited by 1 mM ouabain. Strychnine and parachloromercuriphenylsulfonate (pCS) (both at 1 mM) also strongly inhibited uptake of [³H]GABA, but metabolic inhibitors (2,4-dinitrophenol, potassium cyanide, and malonate) were without effect. Only three structural analogs of GABA (nipecotate, -alanine, and 2,4-diaminobutyrate) inhibited uptake of [³H]GABA, while several other compounds with structural similarities to GABA (e.g. glycine,l-proline, and taurine) did not interact with the system. The kinetic studies indicated presence of a second uptake (K _m=92 M,V _max=0.124 nmol/mg/min) in the primary cultures containing predominantly glioblasts. On the other hand, only one of the neuronal cell lines transformed by simian virus SV40 appeared to accumulate [³H]GABA against a concentration gradient. ApparentK _m of this uptake was relatively high (819 M), and it was only weakly inhibited by 1 mM ouabain and 1 mM pCS. The structural specificity also differed from that of the uptake observed in the primary cultures. Significantly, none of the nontransformed continuous cell lines of either tumoral (glioma, C₆; neuroblastoma, Ml; MINN) or normal (NN; I₆) origin actively accumulated [³H]GABA. It is suggested that for the neurochemical studies related to GABA and requiring homogeneous cell populations, the primary cultures offer a better experimental model than the continuous cell lines.     

Untersuchungen über das Vorkommen der Laufkäfer [Col.: Carabidae] auf Europäischen Getreidefeldern

Zusammenfassung Mit Bodenfallen wurde das Auftreten der Laufk?fer vorwiegend in Winterweizenfeldern in der Zeit von Ende Mai bis Anfang August 1973 und 1974 in Belgien, Deutschland, den Niederlanden und Schweden festgestellt. Unter den h?ufigsten Arten tratenPterostichus vulgaris undBembidion lampros in allen,Agonum dorsale in fast allen Untersuchungsgebieten gemeinsam auf. Sechzehn weitere Arten konnten als allgemein für die meisten Weizenfelder der Untersuchungsgebiete genannt werden. In fast allen Gebieten waren (gemessen an der Artenzahl) die Frühlingstiere mit Herbstbestand in der überzahl Die durchschnittliche Anazahl der pro Falle und Tag gefangenen Laufk?fer variierte von 1 bis 13 und lag im Mittel aller Gebiete bei 6 K?fern. Mit der Quadratmenthode wurden in einem norddeutschen Gebiet 6000 bis 11000 Individuen der ArtAgonum dorsale pro ha festgestellt. Es wird darauf hingewiesen, dass es lohnenswert ist, die Laufk?fer bei der Erstellung integrierter Pflanzenschutzprogramme zu berücksichtigen.

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Summary The occurrence ofCarabidae was investigated by pitfall trapping mainly on fields of winter wheat from the end of May to the beginning of August 1973 and 1974 in Belgium, Germany, the Netherlands, and Sweden. Among the most common speciesPterostichus vulgaris andBembidion lampros were found in all,Agonum dorsale in nearly all regions. Furthermore the following 16 species could be regarded as common for most of the wheat fields in the regions of investigation:Trechus quadristriatus, T. discus, Pterostichus cupreus, P. niger, Asaphidion flavipes, Clivina fossor, Loricera pilicornis, Nebria brevicollis, Harpalus pubescens, H. aeneus, Agonum mülleri, Bembidion ustulatum, Amara familiaris, Calathus fuscipes, Carabus cancellatus ardC. granulatus. In most of the regions, if numbers of species were regarded, spring breeders with an autumn generation were more numerous than spring breedens without an autumn generation, and autumn breeders. The average number of ground beetles per pitfall trap and day varied from 1 to 13 and averaged 6, if all regions were regarded. By using the square method in a Northern German region there was stated a number of 6,000 up to 11,000 individuals ofAgonum dorsale per hectare. Where a comparison was possible, no distinct difference could be stated concerning the number of carabid species between the present investigations and those having taken place ca. 20 years ago. It is emphasized, that it is worth while taking into account the ground beetles, too, when establishing integrated control programs in cereal growing.

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West-Pal?arktisch Regionale Sektion der O.I.L.B., Arbeitsgruppe Integrierte Bek?mpfung im Getreidebau

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Dem Andenken unseres Freundes und KollegenThyge Thygesen (1923–1974) gewidmet, deran der Gründung der Arbeitsgruppe beteiligt vor Abschluss der Untersuchungen verstorben ist.     

Morphological and histochemical evidence of mitoribosomes in Manduca sexta.

The mitochondria found in the neurons of the frontal ganglion of Manduca sexta contained numerous mitoribosomes. The mitochondria of the glial and perineural cells did not contain mitoribosomes. The mitoribosomes were digested in RNase whereas phospholipase C digested the cellular membranes but had no effect on the mitoribosomes.     

Molecular and morphometric analyses identify new lineages within a large Eumida (Annelida) species complex

We report on two new lineages of the Eumida sanguinea complex from Great Britain and describe one of them as a new species using a multilocus approach, including the mitochondrial DNA COI-5P and the nuclear markers ITS (ITS1, 5.8S rRNA and ITS2) and 28S rRNA. The molecular analysis placed Eumida mackiei sp. nov. in a monophyletic clade with 19.1% (COI), 10.1% (ITS) and 1.7% (28S) mean distance to its nearest neighbour. Molecular diagnoses were also applied to nine lineages within the E. sanguinea complex. This was complemented with morphometric data employing multivariate statistical analysis and the incorporation of statistical dissimilarities against three other described species from the complex. Eumida mackiei sp. nov. can be distinguished from E. notata and E. maia by the larger distance between the eyes and differences in morphometric proportions mainly in the dorsal and ventral cirri as well as in the prostomial appendages. E. sanguinea sensu stricto failed to produce a cluster of its own in the morphometric analysis, probably due to juvenile bias. Integrative taxonomy provided strong evidence to formally describe a new cryptic species that can now be used in biomonitoring or other relevant ecological research.     

Sea trout Salmo trutta in the subarctic: home-bound but large variation in migratory behaviour between and within populations

Anadromous brown trout (sea trout), Salmo trutta, is currently in decline throughout its range, largely due to anthropogenic stressors in freshwater and marine habitats. Acoustic telmetry was utilized to study the marine migration of sea trout post-smolts from three populations in a relatively pristine subarctic fjord system. While at sea, the sea trout spent a substantial part of their time close to their natal river, preferred near shore over pelagic habitats and were strongly surface oriented. Despite a fidelity towards local areas, the sea trout utilized various parts of the fjord system, with maximum dispersion >30 km and total migration distance >300 km. Almost half of the sea trout (44%) migrated between river outlets, indicating that a metapopulation approach may be appropriate when managing neighbouring sea trout populations at high latitudes. Furthermore, the different populations displayed different migratory behaviours in terms of distance migrated, dispersion from origin and the likelihood of leaving their home area. This variation in migratory behaviour is likely influenced by spatiotemporal differences in habitat quality between sites, indicating that local habitat variations may promote population-specific behavioural responses even in relatively confined fjord systems.     

Near Infrared Optical Projection Tomography for Assessments of β-cell Mass Distribution in Diabetes Research

By adapting OPT to include the capability of imaging in the near infrared (NIR) spectrum, we here illustrate the possibility to image larger bodies of pancreatic tissue, such as the rat pancreas, and to increase the number of channels (cell types) that may be studied in a single specimen. We further describe the implementation of a number of computational tools that provide: 1/ accurate positioning of a specimen''s (in our case the pancreas) centre of mass (COM) at the axis of rotation (AR)²; 2/ improved algorithms for post-alignment tuning which prevents geometric distortions during the tomographic reconstruction² and 3/ a protocol for intensity equalization to increase signal to noise ratios in OPT-based BCM determinations³. In addition, we describe a sample holder that minimizes the risk for unintentional movements of the specimen during image acquisition. Together, these protocols enable assessments of BCM distribution and other features, to be performed throughout the volume of intact pancreata or other organs (e.g. in studies of islet transplantation), with a resolution down to the level of individual islets of Langerhans.     

A Gene Transfer Agent and a Dynamic Repertoire of Secretion Systems Hold the Keys to the Explosive Radiation of the Emerging Pathogen Bartonella

Gene transfer agents (GTAs) randomly transfer short fragments of a bacterial genome. A novel putative GTA was recently discovered in the mouse-infecting bacterium Bartonella grahamii. Although GTAs are widespread in phylogenetically diverse bacteria, their role in evolution is largely unknown. Here, we present a comparative analysis of 16 Bartonella genomes ranging from 1.4 to 2.6 Mb in size, including six novel genomes from Bartonella isolated from a cow, two moose, two dogs, and a kangaroo. A phylogenetic tree inferred from 428 orthologous core genes indicates that the deadly human pathogen B. bacilliformis is related to the ruminant-adapted clade, rather than being the earliest diverging species in the genus as previously thought. A gene flux analysis identified 12 genes for a GTA and a phage-derived origin of replication as the most conserved innovations. These are located in a region of a few hundred kb that also contains 8 insertions of gene clusters for type III, IV, and V secretion systems, and genes for putatively secreted molecules such as cholera-like toxins. The phylogenies indicate a recent transfer of seven genes in the virB gene cluster for a type IV secretion system from a cat-adapted B. henselae to a dog-adapted B. vinsonii strain. We show that the B. henselae GTA is functional and can transfer genes in vitro. We suggest that the maintenance of the GTA is driven by selection to increase the likelihood of horizontal gene transfer and argue that this process is beneficial at the population level, by facilitating adaptive evolution of the host-adaptation systems and thereby expansion of the host range size. The process counters gene loss and forces all cells to contribute to the production of the GTA and the secreted molecules. The results advance our understanding of the role that GTAs play for the evolution of bacterial genomes.     

An evaluation of analysis pipelines for DNA methylation profiling using the Illumina HumanMethylation450 BeadChip platform

The proper identification of differentially methylated CpGs is central in most epigenetic studies. The Illumina HumanMethylation450 BeadChip is widely used to quantify DNA methylation; nevertheless, the design of an appropriate analysis pipeline faces severe challenges due to the convolution of biological and technical variability and the presence of a signal bias between Infinium I and II probe design types. Despite recent attempts to investigate how to analyze DNA methylation data with such an array design, it has not been possible to perform a comprehensive comparison between different bioinformatics pipelines due to the lack of appropriate data sets having both large sample size and sufficient number of technical replicates. Here we perform such a comparative analysis, targeting the problems of reducing the technical variability, eliminating the probe design bias and reducing the batch effect by exploiting two unpublished data sets, which included technical replicates and were profiled for DNA methylation either on peripheral blood, monocytes or muscle biopsies. We evaluated the performance of different analysis pipelines and demonstrated that: (1) it is critical to correct for the probe design type, since the amplitude of the measured methylation change depends on the underlying chemistry; (2) the effect of different normalization schemes is mixed, and the most effective method in our hands were quantile normalization and Beta Mixture Quantile dilation (BMIQ); (3) it is beneficial to correct for batch effects. In conclusion, our comparative analysis using a comprehensive data set suggests an efficient pipeline for proper identification of differentially methylated CpGs using the Illumina 450K arrays.     

[18F]FLT and [18F]FDG PET for Non-invasive Treatment Monitoring of the Nicotinamide Phosphoribosyltransferase Inhibitor APO866 in Human Xenografts

Introduction

APO866 is a new anti-tumor compound inhibiting nicotinamide phosphoribosyltransferase (NAMPT). APO866 has an anti-tumor effect in several pre-clinical tumor models and is currently in several clinical phase II studies. 3′-deoxy-3′-[18F]fluorothymidine ([18F]FLT) is a tracer used to assess cell proliferation in vivo. The aim of this study was non-invasively to study effect of APO866 treatment on [18F]FLT and 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) uptake.

Methods

In vivo uptake of [18F]FLT and [18F]FDG in human ovary cancer xenografts in mice (A2780) was studied at various time points after APO866 treatment. Baseline [18F]FLT or [18F]FDG scans were made before treatment and repeated after 24 hours, 48 hours and 7 days. Tumor volume was followed with computed tomography (CT). Tracer uptake was quantified using small animal PET/CT. One hour after iv injection of tracer, static PET scans were performed. Imaging results were compared with Ki67 immunohistochemistry.

Results

Tumors treated with APO866 had volumes that were 114% (24 h), 128% (48 h) and 130% (Day 7) relative to baseline volumes at Day 0. In the control group tumor volumes were 118% (24 h), 145% (48 h) and 339% (Day 7) relative to baseline volumes Day 0. Tumor volume between the treatment and control group was significantly different at Day 7 (P = 0.001). Compared to baseline, [18F]FLT SUVmax was significantly different at 24 h (P<0.001), 48 h (P<0.001) and Day 7 (P<0.001) in the APO866 group. Compared to baseline, [18F]FDG SUVmax was significantly different at Day 7 (P = 0.005) in the APO866 group.

Conclusions

APO866 treatment caused a significant decrease in [18F]FLT uptake 24 and 48 hours after treatment initiation. The early reductions in tumor cell proliferation preceded decrease in tumor volume. The results show the possibility to use [18F]FLT and [18F]FDG to image treatment effect early following treatment with APO866 in future clinical studies.     

Thermal habitat of adult Atlantic salmon Salmo salar in a warming ocean

The year-round thermal habitat at sea for adult Atlantic salmon Salmo salar (n = 49) from northern Norway was investigated using archival tags over a 10 year study period. During their ocean feeding migration, the fish spent 90% of the time in waters with temperatures from 1.6–8.4°C. Daily mean temperatures ranged from −0.5 to 12.9°C, with daily temperature variation up to 9.6°C. Fish experienced the coldest water during winter (November–March) and the greatest thermal range during the first summer at sea (July–August). Trends in sea-surface temperatures influenced the thermal habitat of salmon during late summer and autumn (August–October), with fish experiencing warmer temperatures in warmer years. This pattern was absent during winter (November–March), when daily mean temperatures ranged from 3.4–5.0°C, in both colder and warmer years. The observations of a constant thermal habitat during winter in both warmer and colder years, may suggest that the ocean distribution of salmon is flexible and that individual migration routes could shift as a response to spatiotemporal alterations of favourable prey fields and ocean temperatures.