Herbicide effects on planktonic systems of different complexity

Bioassays of different complexity were compared with respect to their capability to predict the environmental impact of the herbicide atrazine in aquatic systems. Acute toxicity tests with Daphnia did not yield meaningful results. Sublethal tests with Daphnia (feeding inhibition, reduction of growth and reproduction) were more sensitive, but effective concentrations of atrazine were still rather high (2 mg/L). A relatively complicated artificial food chain system that incorporated direct and indirect effects on Daphnia yielded significant reduction of daphnid population growth at 0.1 mg/L. Enclosure experiments with natural communities were by far the most sensitive tools. Community responses could be measured at concentrations as low as 1 µg/L and 0.1 µg atrazine/L. At the lowest concentration, however, communities recovered after three weeks. We conclude that in complex systems indirect effects can be more important than direct effects, so that, contrary to the conditions in simple tests, non-target organisms may be the better indicators of herbicide stress to natural communities.     

Centriole as microtubule-organizing centers for marginal bands of molluscan erythrocytes

The erythrocytes of blood clams (arcidae) are flattened, elliptical, and nucleated. They contain elliptical marginal bands (MBs) of microtubules, each physically associated with a pair of centrioles marginal bands (MBs) of microtubles, each physically associated with a pair of centrioles (Cohen, W., and I. Nemhauser, 1980, J. Cell Biol., 86:286-291). The MBs were found to be cold labile in living cells, disappearing within 1-2 h at 0 degrees C. After the cells had been rewarmed for 1-2 h, continuous MBs with associated centrioles were once again present. Time-course studies utilizing phase contrast, antitubulin immunofluorescence, and electron microscopy of cytoskeletons prepared during rewarming revealed structural evidence of centriole participation in MB reassembly. At the earliest stage of reassembly, a continuous MB was not present. Instead, relatively short and straight microtubules focused on a pointed centriolar “pole,” and none were present elsewhere in the cytoskeleton. Thin continuous MBs then formed, still pointed in the centriolar region. Subsequently, the MBs regained ellipticity, with their thickness gradually increasing but not reaching that of controls even after several hours of rewarming. At these later time points, microtubules still radiated from the centrioles and joined the MBs some distance away. In the presence of 0.1 mM colchicines, MB reassembly was arrested at the pointed stage. Electron microscopic observations indicate that pericentriolar material is involved in microtubule nucleation in this system, rather than the centriolar triplets directly. The results suggest a model in which the centrioles and associated material nucleate assembly and growth of microtubules in diverging directions around the cell periphery. Microtubules of opposite polarity would then pass each other at the end of the cell distal to the centrioles, with continued elongation eventually closing the MB ellipse behind the centriole pair.     

Comparison of (R)-[3H] Tomoxetine and (R/S)-[3H] Nisoxetine Binding in Rat Brain

Abstract: ( R )-[³H]Tomoxetine is a radioligand that binds to the norepinephrine (NE) uptake site with high affinity but also binds to a second, lower-affinity site. The goal of the present study was to identify the nature of this low-affinity site by comparing the binding properties of ( R )-[³H]tomoxetine with those of ( R/S )-[³H]nisoxetine, a highly selective ligand for the NE uptake site. In homogenate binding studies, both radioligands bound to the NE uptake site with high affinity, whereas ( R )-[³H]tomoxetine also bound to a second, lower-affinity site. The autoradiographic distribution of binding sites for both radioligands is consistent with the known distribution of NE-containing neurons. However, low levels of ( R )-[³H]-tomoxetine binding were seen in the caudate-putamen, globus pallidus, olfactory tubercle, and zona reticulata of the substantia nigra, where ( R/S )-[³H]nisoxetine binding was almost absent. In homogenates of the caudate-putamen, the NE uptake inhibitors desipramine and ( R )-nisoxetine and the serotonin (5-HT) uptake inhibitor citalopram produced biphasic displacement curves. Autoradiographic studies using 10 n M ( R )-nisoxetine to mask the binding of ( R )-[³H]tomoxetine to the NE uptake site produced autoradiograms that were similar to those produced by [³H]citalopram. Therefore, ( R )-[³H]tomoxetine binds to the NE uptake site with high affinity and the 5-HT uptake site with somewhat lower affinity.     

Metabolomics standards initiative: ontology working group work in progress

In this article we present the activities of the Ontology Working Group (OWG) under the Metabolomics Standards Initiative (MSI) umbrella. Our endeavour aims to synergise the work of several communities, where independent activities are underway to develop terminologies and databases for metabolomics investigations. We have joined forces to rise to the challenges associated with interpreting and integrating experimental process and data across disparate sources (software and databases, private and public). Our focus is to support the activities of the other MSI working groups by developing a common semantic framework to enable metabolomics-user communities to consistently annotate the experimental process and to enable meaningful exchange of datasets. Our work is accessible via a public webpage and a draft ontology has been posted under the Open Biological Ontology umbrella. At the very outset, we have agreed to minimize duplications across omics domains through extensive liaisons with other communities under the OBO Foundry. This is work in progress and we welcome new participants willing to volunteer their time and expertise to this open effort. See the MSI Ontology Working Group website for a complete list of members and contributors. Web URL:     

Identification of a novel integrin alphaMbeta2 binding site in CCN1 (CYR61), a matricellular protein expressed in healing wounds and atherosclerotic lesions

CCN1 (cysteine-rich 61) and CCN2 (connective tissue growth factor) are growth factor-inducible immediate-early gene products found in atherosclerotic lesions, restenosed blood vessels, and healing cutaneous wounds. Both CCN proteins have been shown to support cell adhesion and induce cell migration through interaction with integrin receptors. Recently, we have identified integrin alphaMbeta2 as the major adhesion receptor mediating monocyte adhesion to CCN1 and CCN2 and have shown that the alphaMI domain binds specifically to both proteins. In the present study, we demonstrated that activated monocytes adhered to a synthetic peptide (CCN1-H2, SSVKKYRPKYCGS) derived from a conserved region within the CCN1 C-terminal domain, and this process was blocked by the anti-alphaM monoclonal antibody 2LPM19c. Consistently, a glutathione S-transferase (GST) fusion protein containing the alphaMI domain (GST-alphaMI) bound to immobilized CCN1-H2 as well as to the corresponding H2 sequence in CCN2 (CCN2-H2, TSVKTYRAKFCGV). By contrast, a scrambled CCN1-H2 peptide and an 18-residue peptide derived from an adjacent sequence of CCN1-H2 failed to support monocyte adhesion or alphaMI domain binding. To confirm that the CCN1-H2 sequence within the CCN1 protein mediates alphaMbeta2 interaction, we developed an anti-peptide antibody against CCN1-H2 and showed that it specifically blocked GST-alphaMI binding to intact CCN1. Collectively, these results identify the H2 sequence in CCN1 and CCN2 as a novel integrin alphaMbeta2 binding motif that bears no apparent homology to any alphaMbeta2 binding sequence reported to date.     

Characterization of neuropeptide Y Y1-like and Y2-like receptor subtypes in the mouse brain

To characterize receptor subtypes in the mouse, we performed autoradiographic localization and pharmacological characterization studies using the selective radiolabeled agonists, [(125)I]-Leu(31), Pro(34)-PYY and [(125)I]-PYY 3-36. The pharmacology of [(125)I]-Leu(31), Pro(34)-PYY and [(125)I]-PYY 3-36 binding to mouse brain homogenates were consistent with Y1-like and Y2-like receptors, respectively. Using receptor autoradiography, high Y1-like binding was observed in the islands of Calleja and dentate gyrus. [(125)I]-PYY 3-36 binding was highest in the hippocampus, lateral septum, stria terminalis of the thalamus, and compacta and lateralis of the substantia nigra. In addition, there are differences in receptor distribution in mouse brain compared to other species that may translate into different functional roles for the NPY receptors within each species.     

PET-compatible endothelin receptor radioligands: Synthesis and first in vitro and in vivo studies

The expression and function of endothelin (ET) receptors is abnormal in cardiovascular diseases, tumor progression, and tumor metastasis. In this study, we prepared two [¹⁸F]-fluorinated derivatives of the non-peptide ET_A receptor antagonist PD 156707 and evaluated their ET receptor binding potencies. Ex vivo as well as in vivo biodistribution studies in mice were performed, as well as the metabolism of the radiotracer, which was examined by metabolite analysis in mice and rats. All tested derivatives of PD 156707 exhibited potent in vitro pharmacological characteristics with K_i values comparable to that of the lead compound. The biodistribution studies showed a high accumulation of the tracer in bile and intestine. In vivo we were able to show that the visualization of the heart as a major target organ with high ET_AR expression is possible.     

MALDI-TOF MS of phosphatidylethanolamines: different adducts cause different post source decay (PSD) fragment ion spectra

Matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF MS) is increasingly applied to lipids. However, positional acyl chain analysis of lipids by MALDI was so far scarcely described. In this paper, the fragmentation behavior of phosphatidylethanolamine (PE) is investigated by using post-source decay (PSD) MS. In dependence on the investigated adduct, significant differences could be obtained. It will be shown that in particular the negative ion spectra enable the determination of the individual acyl chains as well as their positions (sn-1 or sn-2). Therefore, MALDI-TOF PSD spectra are a real alternative to more sophisticated MS/MS methods.