Multiple actions of anandamide on neonatal rat cultured sensory neurones
The endogenous cannabinoid anandamide (AEA) can modulate the excitability of sensory neurones; this may be of benefit in the treatment of pain disorders. In this study, the whole cell patch clamp technique and fura-2 calcium imaging have been used to investigate the actions of AEA in cultured dorsal root ganglion (DRG) neurones with particular reference to modulation of ion conductances, and activation of TRPV1 receptors. AEA inhibited voltage-activated Ca2+ currents independently of CB1 receptor activation. AEA had dual effects on depolarisation-evoked Ca2+ flux in distinct subpopulations of DRG neurones; the inhibitory effect was CB1 independent, while the enhancing effect was CB1 mediated but pertussis toxin insensitive, indicating activation of multiple signalling pathways of AEA. AEA both inhibited voltage-activated K+ currents and slowed the recovery phase of depolarisation-evoked Ca2+ transients, both effects being insensitive to pertussis toxin treatment. These effects were also attenuated by the PMSF, an inhibitor of AEA hydrolysis, and mimicked by arachidonic acid, a primary metabolite of AEA, suggesting that production of active metabolites contributes to the observed effects of AEA. AEA elicited TRPV1 mediated responses with less efficacy than capsaicin. When applied directly to the intracellular environment of DRG neurones, the ability of AEA to evoke TRPV1 responses was significantly enhanced as compared with extracellular application. In neurones treated chronically with nerve growth factor (NGF), previously shown to sensitise TRPV1 receptors, responses to AEA and capsaicin were affected differentially. Further investigations revealed a cannabinoid receptor mediated inhibition of TRPV1 activity in low LGF neurones, which was disrupted by chronic NGF treatment.