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International audience; Acid­Sensing Ion Channels (ASICs) form a class of neuronal excitatory cation channels gated by extracellular protons. We have isolated several peptide toxins from seaanemone, spider and snake venoms which allowed to successfully explore the role of these channels in physiopathological processes such as pain. Mambalgin (Mamb­1), was used to reveal the role of subtypes of ASIC channels in pain, including inflammatory and neuropathic pain. It acts as a potent and specific inhibitor of ASIC1a­containing channels, which are the major channel subtypes expressed in the central nervous system, and of peripheral ASIC1b­containing channels that are specific of sensory neurons. Mamb­1 shows potent analgesic effects on thermal and mechanical nociception upon different route of injection in experimental animal models of inflammatory and neuropathic pain. The effects can be as strong as morphine but involve opioid­independent pathways, without producing apparent toxicity. Mamb­1 acts as a gating­modifier that binds to the extracellular domain of the channel to block ASICs by a pH sensor­trapping mechanism. We recently used this peptide to explore the involvement of the ASIC1 in a rodent model of acute and chronic migraine induced by systemic injections of isosorbide dinitrate (ISDN), a therapeutically used nitric oxide donor known to produce headache as side­effect in humans. We studied the effects of systemic injections of Mamb­1 on cutaneous allodynia, by testing cephalic and extra­cephalic mechanical sensitivity. A single ISDN injection induced migraine­like acute symptoms, whereas one daily ISDN injections during four days induced chronic effects. Intravenous injection of Mamb­1 or amiloride, inhibited both the acute and the chronic mechanical allodynia. These data using a venom­ derived, peptide specific inhibitor support the involvement of central and peripheral ASIC1­containing channels in pain and highlight the therapeutic potential of blocking these channels

The snake toxin MT7 is a potent and specific allosteric modulator of the human M1 muscarinic receptor (hM1). We previously characterized by mutagenesis experiments the functional determinants of the MT7-hM1 receptor interaction (Fruchart-Gaillard, C., Mourier, G., Marquer, C., Stura, E., Birdsall, N. J., and Servent, D. (2008) Mol. Pharmacol. 74, 1554-1563) and more recently collected evidence indicating that MT7 may bind to a dimeric form of hM1 (Marquer, C., Fruchart-Gaillard, C., Mourier, G., Grandjean, O., Girard, E., le Maire, M., Brown, S., and Servent, D. (2010) Biol. Cell 102, 409-420). To structurally characterize the MT7-hM1 complex, we adopted a strategy combining double mutant cycle experiments and molecular modeling calculations. First, thirty-three ligand-receptor proximities were identified from the analysis of sixty-one double mutant binding affinities. Several toxin residues that are more than 25 angstrom apart still contact the same residues on the receptor. As a consequence, attempts to satisfy all the restraints by docking the toxin onto a single receptor failed. The toxin was then positioned onto two receptors during five independent flexible docking simulations. The different possible ligand and receptor extracellular loop conformations were described by performing simulations in explicit solvent. All the docking calculations converged to the same conformation of the MT7-hM1 dimer complex, satisfying the experimental restraints and in which (i) the toxin interacts with the extracellular side of the receptor, (ii) the tips of MT7 loops II and III contact one hM1 protomer, whereas the tip of loop I binds to the other protomer, and (iii) the hM1 dimeric interface involves the transmembrane helices TM6 and TM7. These results structurally support the high affinity and selectivity of the MT7-hM1 interaction and highlight the atypical mode of interaction of this allosteric ligand on its G protein-coupled receptor target.