Anandamide transport inhibitor AM404 and structurally related compounds inhibit synaptic transmission between rat hippocampal neurons in culture independent of cannabinoid CB1 receptors

N-(hydroxyphenyl)-arachidonamide (AM404) is an inhibitor of endocannabinoid transport. We examined the effects of AM404 on glutamatergic synaptic transmission using network-driven increases in intracellular Ca2+ concentration ([Ca2+] spikes) as an assay. At a concentration of I muM AM404 inhibited [Ca2+](i) spiking by 73 +/- 8%. The cannabinoid CB I receptor antagonist N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide hydrochloride (SR141716A), the vanilloid VR1 receptor antagonist capsazepine (CPZ), and treatment with pertussis toxin failed to block AM404-mediated inhibition. AM404 (3 muM) inhibited action-potential-evoked Ca2+ influx by 58 +/- 3% but failed to affect calcium influx evoked by depolarization with 30 mM K+ suggesting that the inhibition of electrically evoked [Ca2+](i) increases and that [Ca2+](i) spiking was due to inhibition of Na+ channels. Palmitoylethanolamide (PMEA), capsaicin (CAP) and (5Z,8Z,11Z,14Z)-N-(4-hydroxy-2-methylphenyl)-5,8,11,14-eicosatetraenamide (VDM11), compounds structurally similar to AM404, inhibited [Ca2+](i) spiking by 34 +/- 10%, 42 +/- 18% and 67 +/- 12%, respectively. Thus, AM404 and related compounds inhibit depolarization-induced Ca2+ influx independent of cannabinoid receptors, suggesting caution when using these agents as pharmacological probes to study synaptic transmission. (C) 2004 Elsevier B.V. All rights reserved.