title: Mechanisms of pruritogen-induced activation of itch nerves in isolated mouse skin.
abstract: Chloroquine (CQ) stimulates itch nerves and causes intense scratching in mice by activating the G-protein coupled receptor (GPCR) MrgprA3; it is not known how stimulation of MrgprA3 (or other GPCRs) leads to activation of the itch nerve terminals in the skin, but previous studies have found that transient receptor potential A1 (TRPA1) gene deletion blocks CQ-induced scratching. In the present study we used a novel dorsal skin-nerve preparation to evaluate mechanisms underlying CQ- and histamine-induced action potential discharge in itch nerve terminals. We found that CQ activation of the nerves requires the beta3 isoform of phospholipase C, but TRPA1 or other TRP channel are not required. Evidence is provided for a role for calcium-activated chloride channels such as TMEM16a in GPCR-activation of itch nerve terminals. The mechanism by which TRP channels participate in pruritogen-induced scratching may involve sites of action other than the primary afferent terminals.
  
  
  
  Chloroquine (CQ) and histamine are pruritogens commonly used to study itch in the mouse. A novel skin-nerve preparation was used to evaluate chloroquine (CQ)- and histamine-induced activation of afferent nerves in the dorsal thoracic skin of the mouse. All CQ sensitive nerves were C-fibres, and were also sensitive to histamine. The response to CQ, but not histamine, was largely absent in mrgpr-cluster Δ-/-mice, supporting the hypothesis that CQ evokes itch largely via stimulation of MrgprA3 receptors. The CQ-induced action potential discharge was largely absent in phospholipase Cβ3 knockout animals. The CQ and histamine responses were not influenced by removal of TRPA1, TRPV1, TRPC3 or TRPC6, nor by the TRP channel blocker Ruthenium Red. The bouts of scratching in response to CQ were not different between wild-type and TRPA1-deficient mice. A selective inhibitor of the calcium-activated chloride channel TMEM16A, N-((4-methoxy)-2-naphthyl)-5-nitroanthranilic acid (MONNA), inhibited CQ-induced action potential discharge at itch nerve terminals and bouts of scratching by about 50%. Although TRPA1 and TRPV1 channels may be involved in the scratching responses to intradermal pruritogens, this is unlikely to be due to an effect at the nerve terminals, where chloride channels may play a more important role.
authors.affiliation: Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
authors.forename: F
authors.initials: F
authors.lastname: Ru
authors.affiliation: Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
authors.forename: H
authors.initials: H
authors.lastname: Sun
authors.affiliation: Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.|Department of Pathophysiology, Biomedical Centre Martin, Jessenius Medical School, Comenius University, Martin, Slovakia.
authors.forename: D
authors.identifier: 0000-0002-2210-4930
authors.initials: D
authors.lastname: Jurcakova
authors.affiliation: Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
authors.forename: R A
authors.identifier: 0000-0002-2303-0144
authors.initials: RA
authors.lastname: Herbstsomer
authors.affiliation: Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
authors.forename: J
authors.initials: J
authors.lastname: Meixong
authors.affiliation: Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
authors.forename: X
authors.initials: X
authors.lastname: Dong
authors.affiliation: Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
authors.forename: B J
authors.identifier: 0000-0001-5299-9029
authors.initials: BJ
authors.lastname: Undem
mesh_terms: D000200:Action Potentials; D000818:Animals; D000075369:Anoctamin-1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D002738:Chloroquine / Q000494:pharmacology; D006632:Histamine / Q000494:pharmacology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009475:Neurons, Afferent / Q000378:metabolism / Q000502:physiology*; D054799:Phospholipase C beta / Q000378:metabolism; D011537:Pruritus / Q000378:metabolism* / Q000503:physiopathology; D012867:Skin / Q000187:drug effects / Q000294:innervation*; D000074025:TRPA1 Cation Channel / Q000235:genetics / Q000378:metabolism
chemical_list: C531181:ANO1 protein, mouse; D000075369:Anoctamin-1; D000074025:TRPA1 Cation Channel; D006632:Histamine; D002738:Chloroquine; D054799:Phospholipase C beta
keywords: C-fibers; TRP channels; chemosensitivity; sensory nerves