Anandamide mediates cognitive judgement bias in rats.

In the present study, we investigated the effects of acute pharmacological manipulation of the endocannabinoid (EC) system on the valence of cognitive judgement bias of rats in the ambiguous-cue interpretation (ACI) paradigm. To accomplish this goal, after initial behavioural training, different groups of rats received single, systemic injections of the irreversible anandamide (AEA) hydrolysis inhibitor URB597, the cannabinoid receptor type 1 (CB1) inverse agonist AM251, the cannabinoid receptor type 2 (CB2) inverse agonist AM630, the combination of URB597 and AM251, and a combination of URB597 and AM630 and were subsequently tested with the ACI paradigm. We report that URB597 at a dose of 1 mg/kg significantly biased animals towards positive interpretation of the ambiguous cue and that this effect was abolished by pre-treatment with AM251 (1 mg/kg) or AM630 (1 mg/kg). The CB1 and CB2 inverse agonists administered alone (1 mg/kg) had no statistically significant effects on the interpretation of the ambiguous cue by rats. Our findings suggest involvement of the endocannabinoid system in the mediation of optimistic judgement bias.

Spinal anandamide produces analgesia in neuropathic rats: possible CB(1)- and TRPV1-mediated mechanisms.

The endocannabinoid anandamide (AEA) activates also transient receptor potential vanilloid-1 (TRPV1) channels. However, no data exist on the potential role of spinal TRPV1 activation by AEA in neuropathic pain. We tested the effect of: 1) AEA (5-100 µg), alone or in the presence of an inhibitor of its hydrolysis, and 2) elevated levels of endogenous AEA (following inhibition of AEA hydrolysis), in CCI rats, and the involvement of TRPV1 or cannabinoid CB(1) receptors in the observed effects. Levels of AEA in the spinal cord of CCI rats were measured following all treatments. AEA (50 µg) displayed anti-allodynic and anti-hyperalgesic effects which were abolished by previous antagonism of TRPV1, but not CB(1), receptors. Depending on the administered dose, the selective inhibitor of AEA enzymatic hydrolysis, URB597 (10-100 µg), reduced thermal and tactile nociception via CB(1) or CB(1)/TRPV1 receptors. The anti-nociceptive effects of co-administered per se ineffective doses of AEA (5 µg) and URB597 (5 µg) was abolished by antagonism of CB(1), but not TRPV1, receptors. Spinal AEA levels were increased after CCI, slightly increased further by URB597, 10 µg i.t., and strongly elevated by URB597, 100 µg. Injection of AEA (50 µg) into the lumbar spinal cord led to its dramatic elevation in this tissue, whereas, when a lower dose was used (5 µg) AEA endogenous levels were elevated only in the presence of URB597 (5 µg). We suggest that spinal AEA reduces neuropathic pain via CB(1) or TRPV1, depending on its local concentration.

Immunohistochemical localization of anabolic and catabolic enzymes for anandamide and other putative endovanilloids in the hippocampus and cerebellar cortex of the mouse brain.

An increasing body of evidence indicates that: 1) the endocannabinoid anandamide (AEA) and other unsaturated N-acylethanolamines (NAEs), 2) 12-(S)-lipoxygenase (12-LOX) products of arachidonic acid, and 3) unsaturated N-acyldopamines (NADAs), act as endogenous ligands of transient receptor potential vanilloid type 1 (TRPV1) channels at intracellular binding sites. AEA is synthesized and released "on demand" in neurons from its membrane precursor, N-arachidonoyl-phosphatidylethanolamine, by an N-acyl-phosphatidylethanolamine-specific phospholipase D (NAPE-PLD), and is inactivated by intracellular hydrolysis by fatty acid amide hydrolase (FAAH), whereas catechol-O-methyl-transferase (COMT) was suggested to inactivate NADAs. However, it is not known whether these enzymes or 12-LOX co-localize to any extent with TRPV1 receptors in the brain. In this study we used immunohistochemical techniques (single peroxidase and double immunofluorescence staining), and analyzed the localization of the TRPV1 channel in mouse hippocampal and cerebellar neurons with respect to NAPE-PLD, FAAH, 12-LOX and COMT. Cycloxygenase-2 (COX-2), another putative AEA-degrading enzyme, was also studied. Co-localization between TRPV1 and either NAPE-PLD or FAAH, COX-2, 12-LOX and COMT was found in Ammon's horn (CA3) hippocampal pyramidal neurons and (with the exception of 12-LOX) in some Purkinje cells. At the cellular level, both anabolic and catabolic enzymes appeared as fine grains with immunoperoxidase labeling and were observed in the somatodendritic compartment of CA3 pyramidal cells as well as (with the exception of 12-LOX) in the cytoplasm of Purkinje neurons, in which FAAH and COX-2 immunoreactivities were, however, preferentially localized in the large extension of the dendritic arbor. Our data agree with the hypothesis that, in potential "endovanillergic" neurons, endogenous TRPV1 agonists, and AEA in particular, act as intracellular mediators by being produced from and/or degraded by the same mouse brain cells that express TRPV1 receptors.

Arvanil, anandamide and N-arachidonoyl-dopamine (NADA) inhibit emesis through cannabinoid CB1 and vanilloid TRPV1 receptors in the ferret.

Cannabinoid (CB) agonists suppress nausea and vomiting (emesis). Similarly, transient receptor potential vanilloid-1 (TRPV1) receptor agonists are anti-emetic. Arvanil, N-(3-methoxy-4-hydroxy-benzyl)-arachidonamide, is a synthetic 'hybrid' agonist of CB1 and TRPV1 receptors. Anandamide and N-arachidonoyl-dopamine (NADA) are endogenous agonists at both these receptors. We investigated if arvanil, NADA and anandamide were anti-emetic in the ferret and their mechanism of action. All compounds reduced the episodes of emesis in response to morphine 6 glucuronide. These effects were attenuated by AM251, a CB1 antagonist that was pro-emetic per se, and TRPV1 antagonists iodoresiniferatoxin and AMG 9810, which were without pro-emetic effects. Similar sensitivity to arvanil and NADA was found for prodromal signs of emesis. We analysed the distribution of TRPV1 receptors in the ferret brainstem and, for comparison, the co-localization of CB1 and TRPV1 receptors in the mouse brainstem. TRPV1 immunoreactivity was largely restricted to the nucleus of the solitary tract of the ferret, with faint labeling in the dorsal motor nucleus of the vagus and sparse distribution in the area postrema. A similar distribution of TRPV1, and its extensive co-localization with CB1, was observed in the mouse. Our findings suggest that CB1 and TRPV1 receptors in the brainstem play a major role in the control of emesis by agonists of these two receptors. While there appears to be an endogenous 'tone' of CB1 receptors inhibiting emesis, this does not seem to be the case for TRPV1 receptors, indicating that endogenously released endocannabinoids/endovanilloids inhibit emesis preferentially via CB1 receptors.

Morphine-induced changes in the activity of proopiomelanocortin and prodynorphin systems in zymosan-induced peritonitis in mice.

We have shown that supplementation of proinflammatory agent with a high dose of morphine not only abolishes inflammation-related pain symptoms but also inhibits influx of leukocytes to the inflamed peritoneal cavity. Present investigations focused on effects of morphine on proopiomelanocortin and prodynorphin systems during zymosan-induced peritonitis. Males of SWISS mice were ip injected with zymosan (Z, 40 mg/kg) or zymosan with morphine (ZM, 20 mg/kg). At time 0 (controls) and 4 and 24h after stimulation, peritoneal leukocytes (PTLs) were counted, PTL levels of opioid peptides (beta-endorphin and dynorphin) measured by radioimmunoassays, while mRNAs coding their respective precursors (POMC and PDYN) and receptors (MOR and KOR) determined by QRT-PCR. Influx of inflammatory PTLs, mainly PMNs, was significantly delayed by morphine co-injection. Total levels of beta-endorphin and dynorphin corresponded with PTL numbers, while levels per cell were similar in all groups except of beta-endorphin, decreased in ZM at 4h. Levels of both peptides in peritoneal fluid were increased in Z and ZM groups at 4h, while at 24h only in case of beta-endorphin in Z group. POMC was increased only in ZM group at 4h of peritonitis, while PDYN in both Z and ZM groups at the same time. MOR mRNA was increased 24h after injection in Z and ZM groups, while KOR mRNA was similar in all groups except of decrease in Z at 24h. In conclusion, endogenous opioids and their receptors are involved in zymosan-induced peritonitis and affected in various ways by morphine co-injection.