Inhibition of PaCaMKII-E isoform in the dorsal unpaired median neurosecretory cells of cockroach reduces nicotine- and clothianidin-induced currents

Cellular responses to Ca(2+) require intermediary proteins such as calcium/calmodulin-dependent protein kinase II (CaMKII), which transduces the signal into downstream effects. We recently demonstrated that the cockroach genome encodes five different CaMKII isoforms, and only PaCaMKII-E isoform is specifically expressed in the dorsal unpaired median neurosecretory cells. In the present study, using antisense oligonucleotides, we demonstrated that PaCaMKII-E isoform inhibition reduced nicotine-induced currents through alpha-bungarotoxin-sensitive and -insensitive nicotinic acetylcholine receptor subtypes. Specifically, PaCaMKII-E isoform is sufficient to repress nicotinic current amplitudes as a result of its depression by antisense oligonucleotides. Similar results were found using the neonicotinoid insecticide clothianidin, which acted as a full agonist of dorsal unpaired median neuron nicotinic acetylcholine receptors. Clothianidin current amplitudes are strongly reduced under bath application of PaCaMKII-E antisense oligonucleotides but no significant results are found with alpha-bungarotoxin co-applied, demonstrating that CaMKII-E isoform affects nicotine currents through alpha-bungarotoxin-sensitive and -insensitive receptor subtypes whereas clothianidin currents are reduced via alpha-bungarotoxin-insensitive receptors. In addition, we found that intracellular calcium increase induced by nicotine and clothianidin were reduced by PaCaMKII-E antisense oligonucleotides, demonstrating that intracellular calcium increase induced by nicotine and clothianidin are affected by PaCaMKII-E inhibition. Cellular responses to Ca(2+) require intermediary proteins such as calcium/calmodulin-dependent protein kinase II (CaMKII). We recently demonstrated that the cockroach genome encodes five different CaMKII isoforms and only PaCaMKII-E isoform was specifically expressed in the dorsal unpaired median neurosecretory cells. Here we show that specific inhibition of PaCaMKII-E isoform is associated with a decrease in nicotine- and clothianidin-induced currents. In addition, analysis of calcium changes demonstrates that PaCaMKII-E inhibition induces a decrease in intracellular calcium concentration

Transmembrane potential polarization, calcium influx, and receptor conformational state modulate the sensitivity of the imidacloprid-insensitive neuronal insect nicotinic acetylcholine receptor to neonicotinoid insecticides

Neonicotinoid insecticides act selectively on insect nicotinic acetylcholine receptors (nAChRs). Recent studies revealed that their efficiency was altered by the phosphorylation/dephosphorylation process and the intracellular signaling pathway involved in the regulation of nAChRs. Using whole-cell patch-clamp electrophysiology adapted for dissociated cockroach dorsal unpaired median (DUM) neurons, we demonstrated that intracellular factors involved in the regulation of nAChR function modulated neonicotinoid sensitivity. DUM neurons were known to express two alpha-bungarotoxin-insensitive nAChR subtypes: nAChR1 and nAChR2. Whereas nAChR1 was sensitive to imidacloprid, nAChR2 was insensitive to this insecticide. Here, we demonstrated that, like nicotine, acetamiprid and clothianidin, other types of neonicotinoid insecticides, acted as agonists on the nAChR2 subtype. Using acetamiprid, we revealed that both steady-state depolarization and hyperpolarization affected nAChR2 sensitivity. The measurement of the input membrane resistance indicated that change in the acetamiprid-induced agonist activity was related to the receptor conformational state. Using cadmium chloride, omega-conotoxin GVIA, and (R,S)-(3,4-dihydro-6,7-dimethoxy-isoquinoline-1-yl)-2-phenyl-N,N-di-acetamide (LOE 908), we found that inhibition of calcium influx through high voltage-activated calcium channels and transient receptor potential gamma (TRPgamma) activated by both depolarization and hyperpolarization increased nAChR2 sensitivity to acetamiprid. Finally, using N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (W7), forskolin, and cAMP, we demonstrated that adenylyl cyclase sensitive to the calcium/calmodulin complex regulated internal cAMP concentration, which in turn modulated TRPgamma function and nAChR2 sensitivity to acetamiprid. Similar TRPgamma-induced modulatory effects were also obtained when clothianidin was tested. These findings bring insights into the signaling pathway modulating neonicotinoid efficiency and open novel strategies for optimizing insect pest control

X-ray Linear Dichroism in cubic compounds: the case of Cr3+ in MgAl2O4

The angular dependence (x-ray linear dichroism) of the Cr K pre-edge in MgAl2O4:Cr3+ spinel is measured by means of x-ray absorption near edge structure spectroscopy (XANES) and compared to calculations based on density functional theory (DFT) and ligand field multiplet theory (LFM). We also present an efficient method, based on symmetry considerations, to compute the dichroism of the cubic crystal starting from the dichroism of a single substitutional site. DFT shows that the electric dipole transitions do not contribute to the features visible in the pre-edge and provides a clear vision of the assignment of the 1s-->3d transitions. However, DFT is unable to reproduce quantitatively the angular dependence of the pre-edge, which is, on the other side, well reproduced by LFM calculations. The most relevant factors determining the dichroism of Cr K pre-edge are identified as the site distortion and 3d-3d electronic repulsion. From this combined DFT, LFM approach is concluded that when the pre-edge features are more intense than 4 % of the edge jump, pure quadrupole transitions cannot explain alone the origin of the pre-edge. Finally, the shape of the dichroic signal is more sensitive than the isotropic spectrum to the trigonal distortion of the substitutional site. This suggests the possibility to obtain quantitative information on site distortion from the x-ray linear dichroism by performing angular dependent measurements on single crystals

Nornicotine application on cockroach dorsal unpaired median neurons induces two distinct ionic currents: implications of different nicotinic acetylcholine receptors

The goal of the present study is to examine the agonist action of nornicotine on insect nicotinic acetylcholine receptors. Using patch-clamp techniques on cockroach dorsal unpaired median neurons, we demonstrated that nornicotine induced two distinct ionic currents named types 1 and 2. We found that alpha-bungarotoxin induced a rapid desensitization of type 1 currents whereas type 2 was completely blocked. Interestingly, types 1 and 2 currents were not blocked by the muscarinic antagonist, pirenzepine but by co-application of 1 muM pirenzepine and 0.5 muM alpha-bungarotoxin, suggesting that muscarinic receptors modulated nornicotine-induced current amplitudes. In addition, type 1 current amplitudes were strongly reduced by 20 muM d-tubocurarine and 5 muM mecamylamine which blocked the previously identified alpha-bungarotoxin-insensitive nAChR1 and nAChR2 receptors. Co-application of alpha-bungarotoxin with d-tubocurarine or mecamylamine completely blocked all ionic currents. We propose that types 1 and 2 currents are associated to several nicotinic receptors subtypes, including nAChR1 and nAChR2 receptors. Finally, we conclude that nornicotine could be used as an agonist to identify distinct insect nicotinic receptors

Use of the gLite-WMS in CMS for production and analysis

The CMS experiment at LHC started using the Resource Broker (by the EDG and LCG projects) to submit Monte Carlo production and analysis jobs to distributed computing resources of the WLCG infrastructure over 6 years ago. Since 2006 the gLite Workload Management System (WMS) and Logging \& Bookkeeping (LB) are used. The interaction with the gLite-WMS/LB happens through the CMS production and analysis frameworks, respectively ProdAgent and CRAB, through a common component, BOSSLite. The important improvements recently made in the gLite-WMS/LB as well as in the CMS tools and the intrinsic independence of different WMS/LB instances allow CMS to reach the stability and scalability needed for LHC operations. In particular the use of a multi-threaded approach in BOSSLite allowed to increase the scalability of the systems significantly. In this work we present the operational set up of CMS production and analysis based on the gLite-WMS and the performances obtained in the past data challenges and in the daily Monte Carlo productions and user analysis usage in the experiment

Organisational participation and women - an attitude problem?

Employee participation is a dynamic and contested area of organisational behaviour, attracting continuing academic, practitioner and policy interest and debate. This chapter focuses on organisational participation and women

Gender and Management: new directions in research and continuing patterns in practice

This is the author’s version of the following article. The definitive version is available at www.interscience.wiley.com:Adelina Broadbridge and Jeff Hearn, Gender and management: New directions in research and continuing patterns in practice, 2008, British Journal of Management, (19), s1, 38-49. http://dx.doi.org/10.1111/j.1467-8551.2008.00570.xCopyright: British Academy of Management, Blackwell Publishing Ltdhttp://www.blackwellpublishing.com

RMDAP: A Versatile, Ready-To-Use Toolbox for Multigene Genetic Transformation

Background: The use of transgenes to improve complex traits in crops has challenged current genetic transformation technology for multigene transfer. Therefore, a multigene transformation strategy for use in plant molecular biology and plant genetic breeding is thus needed. Methodology/Principal Findings: Here we describe a versatile, ready-to-use multigene genetic transformation method, named the Recombination-assisted Multifunctional DNA Assembly Platform (RMDAP), which combines many of the useful features of existing plant transformation systems. This platform incorporates three widely-used recombination systems, namely, Gateway technology, in vivo Cre/loxP and recombineering into a highly efficient and reliable approach for gene assembly. RMDAP proposes a strategy for gene stacking and contains a wide range of flexible, modular vectors offering a series of functionally validated genetic elements to manipulate transgene overexpression or gene silencing involved in a metabolic pathway. In particular, the ability to construct a multigene marker-free vector is another attractive feature. The built-in flexibility of original vectors has greatly increased the expansibility and applicability of the system. A proof-ofprinciple experiment was confirmed by successfully transferring several heterologous genes into the plant genome. Conclusions/Significance: This platform is a ready-to-use toolbox for full exploitation of the potential for coordinate regulation of metabolic pathways and molecular breeding, and will eventually achieve the aim of what we call ‘‘one-sto