AChBP-targeted α-conotoxin correlates distinct binding orientations with nAChR subtype selectivity

Neuronal nAChRs are a diverse family of pentameric ion channels with wide distribution throughout cells of the nervous and immune systems. However, the role of specific subtypes in normal and pathological states remains poorly understood due to the lack of selective probes. Here, we used a binding assay based on acetylcholine-binding protein (AChBP), a homolog of the nicotinic acetylcholine ligand-binding domain, to discover a novel α-conotoxin (α-TxIA) in the venom of Conus textile. α-TxIA bound with high affinity to AChBPs from different species and selectively targeted the α3β2 nAChR subtype. A co-crystal structure of Ac-AChBP with the enhanced potency analog TxIA(A10L), revealed a 20° backbone tilt compared to other AChBP–conotoxin complexes. This reorientation was coordinated by a key salt bridge formed between Arg5 (TxIA) and Asp195 (Ac-AChBP). Mutagenesis studies, biochemical assays and electrophysiological recordings directly correlated the interactions observed in the co-crystal structure to binding affinity at AChBP and different nAChR subtypes. Together, these results establish a new pharmacophore for the design of novel subtype-selective ligands with therapeutic potential in nAChR-related diseases

Characterization of a novel alpha-conotoxin TxID from Conus textile that potently blocks rat alpha3/beta4 nicotinic acetylcholine receptors

The alpha 3 beta 4 nAChRs are implicated in pain sensation in the PNS and addiction to nicotine in the CNS. We identified an alpha-4/6-conotoxin (CTx) TxID from Conus textile. The new toxin consists of 15 amino acid residues with two disulfide bonds. TxID was synthesized using solid phase methods, and the synthetic peptide was functionally tested on nAChRs heterologously expressed in Xenopus laevis oocytes. TxID blocked rat alpha 3 beta 4 nAChRs with a 12.5 nM IC50, which places it among the most potent alpha 3 beta 4 nAChR antagonists. TxID also blocked the closely related alpha 6/alpha 3 beta 4 with a 94 nM IC50 but showed little activity on other nAChR subtypes. NMR analysis showed that two major structural isomers exist in solution, one of which adopts a regular alpha-CTx fold but with different surface charge distribution to other 4/6 family members. alpha-CTx TxID is a novel tool with which to probe the structure and function of alpha 3 beta 4 nAChRs

Community informatics for youth: Using new digital media to foster personal growth and community action

During fall 2007 a new collaboration between the Graduate School of Library and Information Science at the University of Illinois Urbana-Champaign and the Extension 4-H network was initiated. Funded by the U.S. Institute of Museum and Library Services, the goal of this collaboration, titled the Youth Community Informatics (YCI) project, is to develop and integrate new and existing strategies to involve youth ages 11-18 from underserved communities in using information and communications technologies to meet community goals. By involving youth from these communities, it is reasoned a sustainable program can be developed through the provisioning of new generations with the knowledge they need to face an information-oriented society. The project is modelled on the work of John Dewey and the American Pragmatists and seeks to create communities of inquiry that bring together professionals, pre-professionals, community youth leaders, and community youth to address community goals in underserved communities. This paper will review the early development of an YCI pedagogy and inquiry units as a means of sharing the curriculum, in addition to highlighting the lessons learned from the first year of this project.published or submitted for publicationis peer reviewe

The effect of peripherally administered cdp-choline in an acute inflammatory pain model: The role of α7 nicotinic acetylcholine receptor

BACKGROUND: CDP-choline (citicholine; cytidine-5'-diphosphate choline) is an endogenously produced nucleotide which, when injected intracerebroventricularly, exerts an antinociceptive effect in acute pain models mediated by central cholinergic mechanisms and alpha 7 nicotinic acetylcholine receptors (alpha 7nAChR). Previous reports also suggest that the peripheral cholinergic system has an antiinflammatory role mediated by alpha 7nAChRs on macrophages. METHODS: We used male Sprague-Dawley rats to assess the antihypersensitivity and antiinflammatory effect of CDP-choline after intraplantar injection of carrageenan (100 mu L, 2%). Mechanical paw withdrawal thresholds and paw thickness were measured by Randall-Selitto testing and microcallipers, respectively. All drugs were administered intraplantarly in a volume 50 mu L. RESULTS: CDP-choline (1, 2.5, 5 mu mol; intraplantar) increased the mechanical paw withdrawal threshold and decreased paw edema in a dose- and time-dependent manner in the carrageenan-injected hindpaw. CDP-choline administration to the noninflamed contralateral hindpaw did not alter ipsdateral inflammation. Methyllycaconitine (100 nmol), a selective alpha 7nAChR antagonist, completely blocked the effects of CDP-choline when administered to the inflamed hindpaw. However, the administration of methyllycaconitine to the contralateral hindpaw did not block the effects of CDP-choline in the ipsilateral paw. The administration of CDP-choline (5 mu mol) 10 min after carrageenan administration to the ipsilateral hindpaw did not reduce swelling and edema but did significantly reduce hypersensitivity. Treatment with CDP-choline decreased tumor necrosis factor-a production in the rat paw tissue after carrageenan. CONCLUSIONS: The results of this study suggest that intraplantar CDP-choline has antihypersensitivity and antiinflammatory effects mediated via alpha 7nAChRs in the carrageenan-induced inflammatory pain model.United States Department of Health & Human Services National Institutes of Health (NIH) - USA (NS048158)United States Department of Health & Human Services National Institutes of Health (NIH) - USA (GM48085)Fulbright Commission TurkeyUnited States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Institute of General Medical Sciences (NIGMS) (R01GM048085)United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Institute of General Medical Sciences (NIGMS) (R37GM048085

A novel mechanism of inhibition of high-voltage activated calcium channels by a-conotoxins contributes to relief of nerve injury-induced neuropathic pain

alpha-Conotoxins that are thought to act as antagonists of nicotinic acetylcholine receptors (nAChRs) containing alpha 3-subunits are efficacious in several preclinical models of chronic pain. Potent interactions of Vc1.1 with other targets have suggested that the pain-relieving actions of alpha-conotoxins might be mediated by either alpha 9 alpha 10 nAChRs or a novel GABA(B) receptor-mediated inhibition of N-type calcium channels. Here we establish that three alpha-conotoxins, Vc1.1, AuIB and MII have distinct selectivity profiles for these three potential targets. Their potencies after intramuscular administration were then determined for reversal of allodynia produced by partial nerve ligation in rats. Vc1.1, which potently inhibits alpha 9 alpha 10 nAChRs and GABA(B)/Ca(2+) channels but weakly blocks alpha 3 beta 2 and alpha 3 beta 4 nAChRs, produced potent, long-lasting reversal of allodynia that were prevented by pre-treatment with the GABA(B) receptor antagonist, SCH50911. alpha-Conotoxin AuIB, a weak alpha 3 beta 4 nAChR antagonist, inhibited GABA(B)/Ca(2+) channels but did not act on alpha 9 alpha 10 nAChRs. AuIB also produced reversal of allodynia. These findings suggest that GABA(B) receptor-dependent inhibition of N-type Ca(2+) channels can mediate the sustained anti-allodynic actions of some alpha-conotoxins. However, MII, a potent alpha 3 beta 2 nAChR antagonist but inactive on alpha 9 alpha 10 and alpha 3 beta 4 nAChRs and GABA(B)/Ca(2+) channels, was demonstrated to have short-acting anti-allodynic action. This suggests that alpha 3 beta 2 nAChRs may also contribute to reversal of allodynia. Together, these findings suggest that inhibition of alpha 9 alpha 10 nAChR is neither necessary nor sufficient for relief of allodynia and establish that alpha-conotoxins selective for GABA(B) receptor-dependent inhibition of N-type Ca(2+) channels relieve allodynia, and could therefore be developed to manage chronic pain. (C) 2010 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved

Conotoxins that Confer Therapeutic Possibilities

Cone snails produce a distinctive repertoire of venom peptides that are used both as a defense mechanism and also to facilitate the immobilization and digestion of prey. These peptides target a wide variety of voltage- and ligand-gated ion channels, which make them an invaluable resource for studying the properties of these ion channels in normal and diseased states, as well as being a collection of compounds of potential pharmacological use in their own right. Examples include the United States Food and Drug Administration (FDA) approved pharmaceutical drug, Ziconotide (Prialt<sup>®</sup>; Elan Pharmaceuticals, Inc.) that is the synthetic equivalent of the naturally occurring ω-conotoxin MVIIA, whilst several other conotoxins are currently being used as standard research tools and screened as potential therapeutic drugs in pre-clinical or clinical trials. These developments highlight the importance of driving conotoxin-related research. A PubMed query from 1 January 2007 to 31 August 2011 combined with hand-curation of the retrieved articles allowed for the collation of 98 recently identified conotoxins with therapeutic potential which are selectively discussed in this review. Protein sequence similarity analysis tentatively assigned uncharacterized conotoxins to predicted functional classes. Furthermore, conotoxin therapeutic potential for neurodegenerative disorders (NDD) was also inferred