Sylvie DIOCHOT, « Ion Channels and Pain » IPMC, CNRS UMR6097, Sophia-Antipolis,Valbonne, France Conflict of interest: none Latrodectism : neurotoxic araneism Intense long lasting abdominal and back pain (muscle cramps) Painful facial « Trismus » from Marectic, 1983 Orlava et al, Nat Struct Biol, 2000 Volynski et al, J Biol Chem 2000 Venom is a complex secretion of animal origin that is use as a defense against an agressor or attack on a prey. Venom can be injected, secreted or sprayed Functions: DEFENSE: Kill PREDATION: Paralyze rapid and effective effects Venoms contains up to 500 toxins Small molécules (MW<1kDa): serotonin, histamin, ATP, alcaloids, glutamate… Polyamines NHH HO O OH O NH CONH2 O NH NH NH HN H2N H NH2 NH Peptides et proteins, Enzymes: PLA2, hyaluronidase, proteases, sphingomyelinase… Public health: Serotherapy Immuno-diagnostic Toxins as structural models for therapeutic molecular design Cancer, coagulation, hypertension and pain therapy Agrifood Insect-Toxins: specific activity on insects, natural insecticide Basic Research Toxins: tools to study membrane receptors and dissect molecular mechanisms of cell signaling. HEMOTOXIC CARDIOTOXIC NEUROTOXIC HYPOTENSIVE VENOMS NECROSIS MYOTOXIC Receptors: - serotonin -Neurotensin - P2X3 … Ion channels activated by: • membrane voltage • membrane strech • ligand binding (NT, peptide, H+…) Technics to study venom toxins effects ELECTROPHYSIOLOGY (Patch clamp) PEPTIDE SYNTHESIS, CLONING MASS SPECTROMETRY PROTEIN SEQUENCING BEHAVIOR TESTS on rodents BINDING STUDIES Radiolabelled toxins CHROMATOGRAPHY -HPLC Crude Venom Ion flux/ion channels Electrical signal Physiological response Chemical signal Hyperexcitability Nav activation Membrane depolarization Cav activation Neurotransmitter release W897X Nav1.7 Mutations –loss of function- Congenital pain insensitivity Mutation –gain of function- Erythromelalgia Nav1.7 I848T (hyperexcitability of nociceptive neurons) Paroxysmal extreme pain disorder Centruroides vittatus Nav1.7 Paw licking after s.c. injection in mice Rowe et al, Plos One, 2011 PAIN ASIC1a ASIC1a/2a Micrurus tener 1b + MitTx Bohlen et al, Nature, 2011 Paw licking after MitTx injection Chinese Traditional medicine Naja kaouthia a-CobraToxin CTX: Presynaptic a7-nAchR antagonist , preventing activation by Ach Chen et al, ActaPharm Sin, 2006 Liu et al, Acta Pharm Sin, 2009 Ziconotide (PRIALT) Conus magus w-ConoTx-MVIIA Severe chronic pain treatment , intrathecal administration Cav2.2 inhibition in sensory neurons Prevent subst P and glutamate release Adverse effects: dizziness, ataxia, confusion, hallucinations, vertigo…reduced therapeutic window Olivera et al, Science, 1985 Schmidtko, The Lancet 2010 Miljanich, Curr Med Chem 2004 Conus marmoreus Trixopelma pruriens m-ConoTx ProTx m-SLPTX-Ssm6a (Peptide 46aa) McIntosh et al, JBC,1995 Bulaj et al, Biochemistry 2006 Yang et al , PNAS, 2013 Hackel et al , PNAS, 2012 Nav1.9 Nav1.7 Nav1.8 ANALGESIA Anthopleura elegantissima Dendroaspis polylepis Psalmopoeus cambridgei Mambalgin APETx2 PcTx1 ASIC1b ASIC1a/2a ANALGESIA Deval et al, EMBO J, 2008 Deval et al, J Neurosci, 2011 ANALGESIA Diochot et al, Nature, 2012 Mazzuca et al, Nature Neurosci, 2007 Geolycosa sp. Purotoxin-1 = peptide (35 aa), PTx Inhibits P2X3 currents (affinity IC50= 12 nM) PTx (2µg) Intraplantar Injections : analgesic effect on rat thermal hyperalgesia model. Grishin et al, Annals of Neurol, 2010 Conus geographus Contulakin-G = 16 aa peptide, « sluggish peptide » Homologies with neurotensin peptides family neurotensin receptor activation (hNTR1, rNTR1&2, mNTR3) Central Injections (i.t., rat, dog) : analgesic effects on inflammatory and thermal pain Absence of adverse effects (motor, cardiac rythm, arterial pressure) CGX-1160 ,Cognetix Inc, Clinique phase II (US. FDA grant in 2006, neuropatic pain) Craig et al, J Biol Chem, 1999 a-Tx APHC1 m-SLPTX Diochot S. 2014
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