nAChRs. A breakthrough was the discovery of endogenous “prototoxin “ Lynx1, belonging to the same family of three- finger proteins as snake neurotoxins, attached to the membranes via glycosyl phosphatidylinositol (GPI) anchor near nAChRs and modulating their function. The report covers our recent findings, including those in collaboration with European laboratories. Methods: HPLC analysis of snake venoms, primary structure determination, NMR and X-ray for 3D-structures, E. coli heterologous expression of three-finger proteins , solid-phase synthesis of alpha-conotoxins. Computer modeling, molecular dynamics , X-ray analysis, electro- physiology and radioligand analysis for theoretical and experimental studies of alpha-conotoxins and other complexes with AChBPs and nAChRs. Results: Novel alpha-conotoxin PnIA analogs and radi- oiodinated derivatives having higher affinity for alpha7 nAChRs and L. stagnalis AChBP were designed and synthe- sized (Kasheverov et al, Mar. Drugs, 2011). Three-finger proteins with an additional disulfide in the loop I, namely WTX Naja kaouthia and water-soluble domain of Lynx1 (ws-lynx1) were expressed in E. coli and shown to act on nAChRs and muscarinic receptors; ws-lynx1 was demon- strated to bind at the classical site for agonists/competitive antagonists at AChBPs and muscle-type nAChRs, but beyond it at neuronal nAChRs (Luykmanova et al, Bio- khimia, 2009; J. Biol. Chem, 2011; Mordvintsev et al, FEBS. J. 2009). A covalent dimer of alpha-cobratoxin, interacting with alpha3beta2 nAChRs, was discovered in Naja kaouthia venom and its X-ray structure recently determined (Osipov et al, J. Biol. Chem. 2008, 2012). Discussion: We demonstrated the advantages of combined approach utilizing novel snake venom protein neurotoxins and synthetic alpha-conotoxins and their analogs as tools in nAChR research. These instruments are supplemented by theoretical and experimental approaches revealing the structure of AChBP complexes which provides information about the binding sites in distinct nAChR subtypes. In particular, with our participation it was demonstrated that binding of alpha-conotoxins (antago- nists) produces conformational changes opposite to those induced by agonists (Celie et al, Nature Str. & Mol. Biol., 2005), while for such antagonists as d-tubocurarine and strychnine different orientations are possible within the same AChBP molecule (Brams et al, PloS Biology, 2011). Conclusion: Peptide and polypeptide neurotoxins provide valuable information about the binding sites in distinct muscle-type and neuronal nAChRs shedding light on functional mechanisms and providing basis for drug design. Keywords: three-finger toxins, alpha-conotoxins, nicotinic acetylcholine receptors 10.1016/j.toxicon.2012.04.143 143. Exploring ‘Labyrinth’ of Pain with Scorpion ‘Sting’ Liu Zhirui, Ji Yonghua Laboratory of Neuropharmacology and Neurotoxicology, Shanghai University, Shanghai, China E-mail address: yhji@staff.shu.edu.cn (J. Yonghua). Background: Voltage-gated sodium channels (VGSCs), responsible for initiation and probagation of action poten- tials in most excitable cells, are implicated to play a critical role in the development and maintenance of pain observed in primary afferent neurons following nerve and tissue injury. Scorpion venemation is a severe medical problem in many regions across the world. Victims of envenoming by a scorpion suffer a variety of pathologies, such as fever, convulsion and intense pain. Discussion: Although the common mechanisms of this pathological pain symptoms have been deduced in part over the years, detail information like pain transmission, integral processing and therapeutic intervention and control have not yet been fully worked out. Currently, it has been demonstrated that the toxic components in scorpion venom are mainly neurotoxic peptides targeting on various ion channels including VGSCs at large. Conclusions: This review will present a state of the art progress in studying the underlying behavioral phenotypes and peripheral mechanisms of pain responses mimicing by a novel experimental BmK (Buthus martensi Karsch) scor- pion sting pain model. Through looking in-depth explora- tion of venom composition, pharmacological binding characteristics, intra-/extra- cellular electrophysiological activities and molecular determinants, a possible cross-talk network regarding the VGSCs-mediated pain initiation and maintanence in overall neuronal sensing and responses pathway by scorpion sting could be approached. Keywords: pain, scorpion sting, BmK, voltage-gated sodium channels 10.1016/j.toxicon.2012.04.144 144. Analgesic Effect of Crotalphine in a New Model of Rat Bone Cancer Pain Yara Cury 1 , Vanessa P. Gutierrez 1 , Patricia Brigatte 2 , Vanessa O. Zambelli 1 , Gisele Picolo 1 , Juliana S. de Carvalho 1 , Fabio Marques 3 1 Laboratorio Especial de Dor e Sinalizacao, Instituto Butantan, Sao Paulo, Brazil 2 Universidade Estadual de Sao Paulo (UNESP), Rio Claro, Brazil 3 Hospital das Clinicas, Universidade de Sao Paulo, Sao Paulo, Brazil E-mail address: yarac@butantan.gov.br (Y. Cury). Background: Crotalphine (CRP), a peptide first identi- fied and isolated from the South American rattlesnake Crotalus durissus terrificus venom, induces analgesic effect mediated by opioid receptors. The aim of this work is to characterize the analgesic effect of crotalphine in a new model of bone cancer pain induced by inoculation of Walker 256 tumor cells into the rat femoral cavity. Methods: Bone tumor implantation and metastasis were determined by histopathological analysis. Bone metabolic alterations were determined by scintigraphy, using 99m Tc-MDP. Femoral images were obtained before and 7, 14 and 21 days after tumor cell injection. Bone cancer pain was characterized by the presence of hyperalgesia (rat paw pressure test) and allodynia (von Frey filaments). Results and Discussion: Photomicrographs analyzed 21 days after injection of tumor cells, demonstrated the presence of tumor cells in the femur of the animals. Abstracts Toxins 2012 / Toxicon 60 (2012) 95–248 168