Inhibition of [3H]batrachotoxinin A-20alpha-benzoate binding to sodium channels and sodium channel function by endocannabinoids

Neurochem Int. 2008 Feb;52(3):438-46. doi: 10.1016/j.neuint.2007.08.004. Epub 2007 Aug 13.


A number of putative endocannabinoids were found to modify the binding of [(3)H]batrachotoxinin A-20alpha-benzoate ([(3)H]BTX-B) to site 2 on voltage-gated sodium channels of mouse brain and achieve functional inhibition of sodium channels in vitro. 2-Arachidonoyl-glycerol (2-AG), arachidonoyl glycerol ether (AGE), N-arachidonoyl-dopamine (NADA) gave almost complete inhibition of [(3)H]BTX-B binding with IC(50) values of 90.4, 51.2 and 20.7 microM, respectively. The CB1 receptor antagonist AM251 (2 microM) had no effect on the displacement of radioligand by these endocanabinoids. Arachidonoyl-glycine (A-Gly) and arachidonoyl-GABA (A-GABA) were apparently less effective inhibitors of [(3)H]BTX-B binding giving 14.8+/-2.2 and 23.9+/-4.8% inhibition at 100 microM. Phenylmethanesulphonylfluoride (PMSF) did not alter the inhibitory effects of 2-AG, AGE, NADA and A-Gly on binding, but the efficacy of 100 microM A-GABA was increased by 60.3+/-6.3% (P<0.05). Scatchard analyses showed that 2-AG, AGE and NADA reduce the binding of [(3)H]BTX-B by lowering B(max) although increases in K(D) were also evident for AGE and NADA. Our kinetic experiments found that 2-AG, AGE and NADA increase the dissociation velocity of radioligand from site 2 on sodium channels demonstrating that these endocannabinoids operate as allosteric inhibitors of [(3)H]BTX-B binding. 2-AG, AGE and NADA inhibited veratridine-dependent (TTX-suppressible) depolarization of the plasma membrane of synaptoneurosomes at low micromolar concentrations and again the capacities of A-Gly and A-GABA to inhibit this response were less pronounced. The three most effective endocannabinoids (2-AG, AGE and NADA) were then examined in a synaptosomal transmitter release assay where they were observed to inhibit sodium channel- (veratridine-dependent) release of l-glutamate and GABA in the low micromolar range. These effects also occurred through a mechanism that was not influenced by 2 microM AM251. It is concluded that direct inhibition of sodium channel function leading to reduced neuronal excitation and depression of presynaptic release of amino acid transmitters is a property shared by several endocannabinoids.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Regulation / drug effects
  • Animals
  • Arachidonic Acids / pharmacology
  • Batrachotoxins / antagonists & inhibitors
  • Batrachotoxins / metabolism*
  • Batrachotoxins / pharmacokinetics
  • Binding Sites / drug effects
  • Binding, Competitive / drug effects*
  • Brain / drug effects*
  • Brain / metabolism
  • Cannabinoid Receptor Modulators / chemistry
  • Cannabinoid Receptor Modulators / metabolism
  • Cannabinoid Receptor Modulators / pharmacology*
  • Cell Membrane / drug effects
  • Cell Membrane / metabolism
  • Dose-Response Relationship, Drug
  • Endocannabinoids*
  • Glycerides / pharmacology
  • Mice
  • Molecular Structure
  • Neurotransmitter Agents / metabolism
  • Pharmacokinetics
  • Presynaptic Terminals / drug effects
  • Presynaptic Terminals / metabolism
  • Radioligand Assay
  • Receptor, Cannabinoid, CB1 / agonists
  • Receptor, Cannabinoid, CB1 / antagonists & inhibitors
  • Receptor, Cannabinoid, CB1 / metabolism
  • Sodium Channels / drug effects*
  • Sodium Channels / metabolism
  • Synaptic Transmission / drug effects*
  • Synaptosomes
  • Tritium


  • Arachidonic Acids
  • Batrachotoxins
  • Cannabinoid Receptor Modulators
  • Endocannabinoids
  • Glycerides
  • Neurotransmitter Agents
  • Receptor, Cannabinoid, CB1
  • Sodium Channels
  • Tritium
  • batrachotoxinin A 20-alpha-benzoate
  • glyceryl 2-arachidonate