title: Neuropathic Nav1.3-mediated sensitization to P2X activation is regulated by protein kinase C.
abstract: BACKGROUND
  Increased neuronal excitability and spontaneous firing are hallmark characteristics of injured sensory neurons. Changes in expression of various voltage-gated Na+ channels (VGSCs) have been observed under neuropathic conditions and there is evidence for the involvement of protein kinase C (PKC) in sensory hyperexcitability. Here we demonstrate the contribution of PKC to P2X-evoked VGSC activation in dorsal root ganglion (DRG) neurons in neuropathic conditions.
  
  
  RESULTS
  Using the spinal nerve ligation (SNL) model of neuropathic pain and whole-cell patch clamp recordings of dissociated DRG neurons, we examined changes in excitability of sensory neurons after nerve injury and observed that P2X3 purinoceptor-mediated currents induced by α,β-meATP triggered activation of TTX-sensitive VGSCs in neuropathic nociceptors only. Treatment of neuropathic DRGs with the PKC blocker staurosporine or calphostin C decreased the α,β-meATP-induced Na+ channels activity and reversed neuronal hypersensitivity. In current clamp mode, α,β-meATP was able to evoke action-potentials more frequently in neuropathic neurons than in controls. Pretreatment with calphostin C significantly decreased the proportion of sensitized neurons that generated action potentials in response to α,β-meATP. Recordings measuring VGSC activity in neuropathic neurons show significant change in amplitude and voltage dependence of sodium currents. In situ hybridization data indicate a dramatic increase in expression of embryonic Nav1.3 channels in neuropathic DRG neurons. In a CHO cell line stably expressing the Nav1.3 subunit, PKC inhibition caused both a significant shift in voltage-dependence of the channel in the depolarizing direction and a decrease in current amplitude.
  
  
  CONCLUSIONS
  Neuropathic injury causes primary sensory neurons to become hyperexcitable to ATP-evoked P2X receptor-mediated depolarization, a phenotypic switch sensitive to PKC modulation and mediated by increased activity of TTX-sensitive VGSCs. Upregulation in VGSC activity after injury is likely mediated by increased expression of the Nav1.3 subunit, and the function of the Nav1.3 channel is regulated by PKC.
authors.affiliation: Montreal Neurological Institute and The Alan Edwards Centre for Research on Pain, Department of Neurology & Neurosurgery, McGill University, Montreal, Canada.
authors.forename: Gary
authors.initials: G
authors.lastname: Mo
authors.forename: Rebecca
authors.initials: R
authors.lastname: Grant
authors.forename: Dajan
authors.initials: D
authors.lastname: O'Donnell
authors.forename: David S
authors.initials: DS
authors.lastname: Ragsdale
authors.forename: Chang-Qing
authors.initials: CQ
authors.lastname: Cao
authors.forename: Philippe
authors.initials: P
authors.lastname: Séguéla
mesh_terms: D000200:Action Potentials / Q000187:drug effects; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D005727:Ganglia, Spinal / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D017403:In Situ Hybridization; D008297:Male; D062552:NAV1.3 Voltage-Gated Sodium Channel; D009281:Naphthalenes / Q000494:pharmacology; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D058469:Receptors, Purinergic P2X / Q000378:metabolism*; D015222:Sodium Channels / Q000378:metabolism*; D019311:Staurosporine / Q000494:pharmacology
chemical_list: D062552:NAV1.3 Voltage-Gated Sodium Channel; D009281:Naphthalenes; D009419:Nerve Tissue Proteins; D058469:Receptors, Purinergic P2X; C478638:Scn3a protein, rat; D015222:Sodium Channels; D000255:Adenosine Triphosphate; D011493:Protein Kinase C; D019311:Staurosporine; C058819:calphostin C; C002630:alpha,beta-methyleneadenosine 5'-triphosphate