Multiple actions of anandamide on neonatal rat cultured sensory neurones

1 We have investigated the effects of the endocannabinoid anandamide ( AEA) on neuronal excitability and vanilloid TRPV1 receptors in neonatal rat cultured dorsal root ganglion neurones. 2 Using whole-cell patch-clamp electrophysiology, we found that AEA inhibits high-voltage-activated Ca2+ currents by 33+/-9% (five out of eight neurones) in the absence of the CB1 receptor antagonist SR141716A (100 nM) and by 32+/-6% (seven out of 10 neurones) in the presence of SR141716A. 3 Fura-2 fluorescence Ca2+ imaging revealed that AEA produced distinct effects on Ca2+ transients produced by depolarisation evoked by 30 mM KCl. In a population of neurones of larger somal area (372+/-20 mum(2)), it significantly enhanced Ca2+ transients (80.26+/-13.12% at 1 muM), an effect that persists after pertussis toxin pretreatment. In a population of neurones of smaller somal area (279+/-18 mum(2)), AEA significantly inhibits Ca2+ transients (30.75+/-3.54% at 1 muM), an effect that is abolished by PTX pretreatment. 4 Extracellular application of 100 nM AEA failed to evoke TRPV1 receptor inward currents in seven out of eight neurones that responded to capsaicin (1 muM), with a mean inward current of -0.94+/-0.21 nA. In contrast, intracellular application of 100 nM AEA elicited robust inward currents in similar to62% of neurones, the mean population response was -0.85+/-0.21 nA. When AEA was applied to the intracellular environment with capsazepine (1 muM), the mean population inward current was -0.01+/-0.01 nA. Under control conditions, mean population current fluctuations of -0.09+/-0.05 nA were observed.