Activation of M3 muscarinic receptors inhibits T-type Ca(2+) channel currents via pertussis toxin-sensitive novel protein kinase C pathway in small dorsal root ganglion neurons

Cell Signal. 2011 Jun;23(6):1057-67. doi: 10.1016/j.cellsig.2011.02.001. Epub 2011 Feb 15.


Cobrotoxin (CbT), a short-chain postsynaptic α-neurotoxin, has been reported to play a role in analgesia. However, to date, the detailed mechanisms still remain unknown. In the present study, we identify a novel functional role of CbT in modulating T-type Ca(2+) channel currents (T-currents) in small dorsal root ganglia (DRG) neurons as well as pain behaviors in mice. We found that CbT inhibited T-currents in a dose-dependent manner. CbT at 1μM reversibly inhibited T-currents by ~26.3%. This inhibitory effect was abolished by the non-selective muscarinic acetylcholine receptor (mAChR) antagonist atropine, or the selective M3 mAChR antagonist 4-DAMP, while naloxone, an opioid receptor antagonist had no effect. Intracellular infusion of GDP-β-S or pretreatment of the cells with pertussis toxin (PTX) completely blocked the inhibitory effects of CbT. Using depolarizing prepulse, we found the absence of direct binding between G-protein βγ subunits and T-type Ca(2+) channels in CbT-induced T-current inhibition. CbT responses were abolished by the phospholipase C inhibitor U73122 (but not the inactive analog U73343). The classical and novel protein kinase C (nPKC) antagonist chelerythrine chlorid or GF109203X abolished CbT responses, whereas the classical PKC antagonist Ro31-8820 or inhibition of PKA elicited no such effects. Intrathecal administration of CbT (5μg/kg) produced antinociceptive effects in mechanical, thermal, and inflammatory pain models. Moreover, CbT-induced antinociception could be abrogated by 4-DAMP. Taken together, these results suggest that CbT acting through M3 mAChR inhibits T-currents via a PTX-sensitive nPKC pathway in small DRG neurons, which could contribute to its analgesic effects in mice.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Calcium Channels, T-Type / metabolism*
  • Calcium Signaling / drug effects
  • Cholera Toxin / pharmacology
  • Cobra Neurotoxin Proteins / pharmacology
  • Estrenes / pharmacology
  • Ganglia, Spinal / cytology*
  • Ganglia, Spinal / metabolism
  • Indoles / pharmacology
  • Maleimides / pharmacology
  • Mice
  • Mice, Inbred ICR
  • Muscarinic Antagonists / pharmacology
  • Neurons / drug effects*
  • Neurons / metabolism
  • Pain Measurement
  • Pertussis Toxin / pharmacology*
  • Piperidines / pharmacology
  • Protein Kinase C / antagonists & inhibitors*
  • Protein Kinase C / metabolism
  • Pyrrolidinones / pharmacology
  • Receptor, Muscarinic M3 / agonists*
  • Type C Phospholipases / antagonists & inhibitors


  • Calcium Channels, T-Type
  • Cobra Neurotoxin Proteins
  • Estrenes
  • Indoles
  • Maleimides
  • Muscarinic Antagonists
  • Piperidines
  • Pyrrolidinones
  • Receptor, Muscarinic M3
  • 1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione
  • 4-diphenylacetoxy-1,1-dimethylpiperidinium
  • Cholera Toxin
  • Pertussis Toxin
  • Protein Kinase C
  • Type C Phospholipases
  • bisindolylmaleimide I