2-phenyl-4-4-5-5-tetramethylimidazoline-1-oxyl-3-oxide and anandamide

Cannabinoid compounds usually produce biphasic effects in the modulation of emotional responses. Low doses of the endocannabinoid anandamide (AEA) injected into the dorsolateral periaqueductal gray matter (dlPAG) induce anxiolytic-like effects via CB1 receptors activation. However, at higher doses the drug loses this effect, in part by activating Transient Receptor Potential Vanilloid Type 1 (TRPV1). Activation of these latter receptors could induce the formation of nitric oxide (NO). Thus, the present study tested the hypothesis that at high doses AEA loses it anxiolytic-like effect by facilitating, probably via TRPV1 receptor activation, the formation of NO. Male Wistar rats received combined injections into the dlPAG of vehicle, the TRPV1 receptor antagonist 6-iodo-nordihydrocapsaicin or the NO scavenger carboxy-PTIO (c-PTIO), followed by vehicle or AEA, and were submitted to the elevated plus maze (EPM) or the Vogel conflict test (VCT). A low dose (5pmol) of AEA produced an anxiolytic-like effect that disappeared at higher doses (50 and 200pmol). The anxiolytic-like effects of these latter doses, however, were restored after pre-treatment with a low and ineffective dose of c-PTIO in both animal models. In addition, the combined administration of ineffective doses of 6-iodo-nordihydrocapsaicin (1nmol) and c-PTIO (0.3nmol) produced an anxiolytic-like response. Therefore, these results support the hypothesis that intra-dlPAG injections of high doses of AEA lose their anxiolytic effects by favoring TRPV1 receptors activity and consequent NO formation, which in turn could facilitate defensive responses.

Topics: Animals; Anxiety; Arachidonic Acids; Cannabinoid Receptor Agonists; Capsaicin; Cyclic N-Oxides; Disease Models, Animal; Drinking; Drug Interactions; Endocannabinoids; Free Radical Scavengers; Imidazoles; Male; Maze Learning; Microinjections; Nitric Oxide; Periaqueductal Gray; Polyunsaturated Alkamides; Rats; Rats, Wistar; Reaction Time; TRPV Cation Channels

The endocannabinoids anandamide (AEA) and 2-arachidonoylglycerol (2-AG) are metabolised by cells by hydrolysis to arachidonic acid followed by esterification into phospholipids. Here, we report that nitric oxide (NO) donors significantly increase the amount of tritium accumulated in the cell membranes of RBL2H3 rat basophilic cells, 3T3-L1 mouse fibroblast cells and b.End5 mouse brain endothelioma cells following incubation of the intact cells with AEA labelled in the arachidonate part of the molecule. Similar results were seen with 2-AG and with arachidonic acid, whilst the NO donors reduced the accumulation of tritium after incubation of RBL2H3 cells with AEA labelled in the ethanolamine part of the molecule. Pretreatment of intact cells with NO donors did not increase the activity of the enzyme mainly responsible for metabolism of AEA, fatty acid amide hydrolase (FAAH). Furthermore, inhibition of FAAH completely blocked the effect produced by NO donors in cells with a large FAAH component, suggesting that for AEA, the effects were downstream of the enzyme. These data raise the possibility that the cellular processing of endocannabinoids following its uptake can be regulated by nitric oxide.

Topics: Amides; Animals; Arachidonic Acids; Benzamides; Cannabinoid Receptor Modulators; Carbamates; Cell Line; Cell Membrane; Cyclic N-Oxides; Endocannabinoids; Ethanolamines; Free Radical Scavengers; Glycerides; Imidazoles; Lipid Metabolism; Mice; Nitric Oxide; Nitric Oxide Donors; Palmitic Acids; Polyunsaturated Alkamides; Rats; Signal Transduction; Tritium