Multiple actions of anandamide on neonatal rat cultured sensory neurones

Abstract

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-voltageactivated Ca 2 þ currents by 3379% (five out of eight neurones) in the absence of the CB 1 receptor antagonist SR141716A (100 nM) and by 3276% (seven out of 10 neurones) in the presence of SR141716A. 3 Fura-2 fluorescence Ca 2 þ imaging revealed that AEA produced distinct effects on Ca 2 þ transients produced by depolarisation evoked by 30 mM KCl. In a population of neurones of larger somal area (372720 mm 2 ), it significantly enhanced Ca 2 þ transients (80.26713.12% at 1 mM), an effect that persists after pertussis toxin pretreatment. In a population of neurones of smaller somal area (279718 mm 2 ), AEA significantly inhibits Ca 2 þ transients (30.7573.54% at 1 mM), 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 mM), with a mean inward current of À0.9470.21 nA. In contrast, intracellular application of 100 nM AEA elicited robust inward currents in B62% of neurones, the mean population response was À0.8570.21 nA. When AEA was applied to the intracellular environment with capsazepine (1 mM), the mean population inward current was À0.0170.01 nA. Under control conditions, mean population current fluctuations of À0.0970.05 nA were observed.

Localization of the Endocannabinoid-Degrading Enzyme Fatty Acid Amide Hydrolase in Rat Dorsal Root Ganglion Cells and Its Regulation after Peripheral Nerve Injury

“…This is consistent with studies demonstrating that antagonism of DRG CB 1 receptors increases the potency of AEA at TRPV1 receptors (Ahluwalia et al, 2003a). At low concentrations, exogenous AEA can reduce TRPV1-evoked excitatory responses via the activation of CB 1 receptors coupled to inhibitory signaling systems (Richardson et al, 1998;Ellington et al, 2002;Ahluwalia et al, 2003b;Evans et al, 2004).…”

“…AEA can activate DRG TRPV1 receptors when applied to the intracellular side of the plasma membrane, with greater potency than on the extracellular side (Hwang et al, 2000;Evans et al, 2004). Considering its lower affinity for TRPV1 than CB 1 receptors (Ahluwalia et al, 2003a,b;Ross, 2003), short half-life, and lipophilic nature, AEA is likely to bind to TRPV1 receptors close to its site of synthesis inside the same neuron.…”

Novel Compounds That Interact with Both Leukotriene B4 Receptors and Vanilloid TRPV1 Receptors

“…The actions of both LTB 4 and LTB 4 ethanolamide are significantly antagonized by the TRPV1 antagonist capsazepine, suggesting that the [Ca 2ϩ ] i response is indeed TRPV1 receptor-mediated. Jung et al (1999, and Jordt and Julius (2002) have published studies indicating that the agonist-binding site of TRPV1 may be intracellular, and anandamide has been reported to be more potent at the TRPV1 receptor when applied intracellularly (Evans et al, 2004), presumably a consequence of the putative intracellular binding location. Conversely, Vyklicky et al (2003) reported that TRPV1 is not activated by vanilloids applied intracellularly, and a study by Chou et al (2004) suggests that the ultrapotent TRPV1 agonist resinaferatoxin interacts with extracellular residues.…”

Anandamide produced by Ca2+-insensitive enzymes induces excitation in primary sensory neurons

“…The concentration of anandamide found in the superfusate (~10 nM) is significantly lower than the concentration of exogenous anandamide needed to induce TRPV1-mediated excitatory effects [1,12,14,38,51]. Therefore, we hypothesised that the excitatory effect of anandamide of PSN origin is mediated through autocrine signalling.…”