arachidonoylserotonin and Disease-Models--Animal

Selective stimulation of carotid chemoreceptors by intravenous infusion of low doses of potassium cyanide (KCN) produces short-lasting escape responses that have been proposed as a model of panic attack. In turn, preclinical studies suggest that facilitation of the endocannabinoid system attenuate panic-like responses. Here, we compared the effects of cannabinoid-related compounds to those of alprazolam, a clinically effective panicolytic, on the duration of the escape reaction induced by intravenous infusion of KCN (80μg) in rats. Alprazolam (1, 2, 4mg/kg) decreased escape duration at doses that did not alter basal locomotor activity. URB597 (0.1, 0.3, 1mg/kg; inhibitor of anandamide hydrolysis), WIN55,212-2 (0.1, 0.3, 1mg/kg; synthetic cannabinoid), arachidonoyl-serotonin (1, 2.5, 5mg/kg; dual TRPV1 and anandamide hydrolysis inhibitor), and cannabidiol (5, 10, 20, 40mg/kg; a phytocannabinoid) did not decrease escape duration. Alprazolam also prevented the increase in arterial pressure evoked by KCN, while bradycardia was unchanged. This study reinforces the validity of the KCN-evoked escape as a model of panic attack. However, it does not support a role for the endocannabinoid system in this behavioral response. These results might have implications for the screening of novel treatments for panic disorder.

Topics: Alprazolam; Analgesics; Animals; Arachidonic Acids; Benzamides; Benzoxazines; Blood Pressure; Cannabinoids; Carbamates; Disease Models, Animal; Dose-Response Relationship, Drug; Enzyme Inhibitors; Escape Reaction; Heart Rate; Hypnotics and Sedatives; Locomotion; Male; Mice; Morpholines; Naphthalenes; Panic Disorder; Potassium Cyanide; Rats, Wistar; Serotonin

There is considerable evidence to support the role of anandamide (AEA), an endogenous ligand of cannabinoid receptors, in neuropathic pain modulation. AEA also produces effects mediated by other biological targets, of which the transient receptor potential vanilloid type 1 (TRPV1) has been the most investigated. Both, inhibition of AEA breakdown by fatty acid amide hydrolase (FAAH) and blockage of TRPV1 have been shown to produce anti-nociceptive effects. Recent research suggests the usefulness of dual-action compounds, which may afford greater anti-allodynic efficacy. Therefore, in the present study, we examined the effect of N-arachidonoyl-serotonin (AA-5-HT), a blocker of FAAH and TRPV1, in a rat model of neuropathic pain after intrathecal administration. We found that treatment with AA-5-HT increased the pain threshold to mechanical and thermal stimuli, with highest effect at the dose of 500nM, which was most strongly attenuated by AM-630, CB2 antagonist, administration. The single action blockers PF-3845 (1000nM, for FAAH) and I-RTX (1nM, for TRPV1) showed lower efficacy than AA-5-HT. Moreover AA-5-HT (500nM) elevated AEA and palmitoylethanolamide (PEA) levels. Among the possible targets of these mediators, only the mRNA levels of CB2, GPR18 and GPR55, which are believed to be novel cannabinoid receptors, were upregulated in the spinal cord and/or DRG of CCI rats. It was previously reported that AA-5-HT acts in CB1 and TRPV1-dependent manner after systemic administration, but here for the first time we show that AA-5-HT action at the spinal level involves CB2, with potential contributions from GRP18 and/or GPR55 receptors.

Topics: Amidohydrolases; Analgesics; Animals; Arachidonic Acids; Cannabinoid Receptor Antagonists; Disease Models, Animal; Dose-Response Relationship, Drug; Endocannabinoids; Ganglia, Spinal; Glycerides; Injections, Spinal; Male; Neuralgia; Nociception; Pain Threshold; Polyunsaturated Alkamides; Rats, Wistar; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2; Receptors, Cannabinoid; Receptors, G-Protein-Coupled; Serotonin; Signal Transduction; Spinal Cord; Time Factors; TRPV Cation Channels

CB(1) receptors in the amygdala have been shown to mediate learned and unlearned anxiety states, however, the role of amygdalar TRPV1 receptors remains unclear. In the present study we investigated the potential anxiolytic action of intra-basolateral amygdala (BLA) infusion of N-arachidonoyl-serotonin (AA-5-HT), a dual blocker of the endocannabinoid-inactivating enzyme, fatty acid amide hydrolase (FAAH), and a TRPV1 antagonist. Varying doses of AA-5-HT (0-0.5 nmol) were administered into the BLA prior to elevated plus maze testing. AA-5-HT significantly increased both time spent and number of entries into the open arms. Next, to determine whether the anxiolytic effects were the result of blocking FAAH, TRPV1, or whether a combined action was required, rats were given intra-BLA infusions of either 0.25 nmol AA-5-HT, 1.0 nmol capsazepine (CZP, a TRPV1 antagonist), 0.01 μg URB597 (a selective FAAH inhibitor), or vehicle. Again, AA-5-HT increased the time spent in the open arms as well as the number of open arm entries. In contrast, CZP and URB597 did not reliably alter plus maze performance. We then investigated the effects of co-administration of CZP (1.0 or 10.0 nmol) and URB597 (0.01 or 0.1 μg). At lower doses, co-injections significantly increased both open arm entries as well as the time spent in the open arms, compared to vehicle or either compound alone. While co-administration of the higher doses had no significant effect when compared to either vehicle or CZP treatment, we did observe that open arm activity was elevated in rats receiving combined CZP-URB597 treatment compared to URB597 alone. Overall, our findings indicate that simultaneous FAAH activity and TRPV1 activation are important with respect to the expression of unconditioned fear as mediated within the BLA.

Topics: Amygdala; Analysis of Variance; Animals; Anxiety Disorders; Arachidonic Acids; Benzamides; Capsaicin; Carbamates; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Exploratory Behavior; Male; Maze Learning; Rats; Rats, Sprague-Dawley; Serotonin; TRPV Cation Channels

The endocannabinoids anandamide and 2-arachidonoylglycerol, and the anandamide-congener, palmitoylethanolamide, are all substrates for the enzyme fatty acid amide hydrolase, and are endowed with anti-inflammatory actions exerted via cannabinoid receptors or, in the case of palmitoylethanolamide, also via other targets. We investigated the role of the endocannabinoid system during granuloma formation, a model of chronic inflammation sustained by neoangiogenesis, in rats. Granuloma was induced by subcutaneous lambda-carrageenin-soaked sponge implants on the back of male Wistar rats. After 96h, granulomas were detached and tissue formation was evaluated as wet weight; the endocannabinoid system was evaluated by the measurement of endocannabinoid levels, by LC-MS, and of cannabinoid receptor expression, by western blot analysis. Moreover, angiogenesis was evaluated by the measurement of both hemoglobin content and CD31 protein expression. Arachidonoylserotonin (AA-5-HT, 12.5-50mug/ml), an inhibitor of FAAH, and palmitoylethanolamide (PEA, 200-800mug/ml) were given locally only once at the time of implantation. Granuloma formation was accompanied by a significant decrease in endocannabinoid and palmitoylethanolamide levels paralleled by increased levels of the fatty acid amide hydrolase, responsible for their breakdown. Moreover, an increase of cannabinoid receptor expression was also observed. Pharmacological elevation of endocannabinoids and palmitoylethanolamide, obtained separately by arachidonoylserotonin and exogenous palmitoylethanolamide treatment, dose-dependently reduced inflammatory hallmarks including tumor necrosis factor-alpha as well as granuloma-dependent angiogenesis. The effect of arachidonoylserotonin was accompanied by near-normalization of 2-arachidonoylglycerol and palmitoylethanolamide levels in the tissue. These findings suggest that chronic inflammation might develop also because of endocannabinoid and palmitoylethanolamide tissue concentration impairment, the correction of which might be exploited to develop new anti-inflammatory drugs.

Topics: Amides; Amidohydrolases; Animals; Arachidonic Acids; Cannabinoid Receptor Modulators; Carrageenan; Chronic Disease; Disease Models, Animal; Dose-Response Relationship, Drug; Endocannabinoids; Ethanolamines; Granuloma; Hemoglobins; Inflammation; Male; Neovascularization, Pathologic; Palmitic Acids; Phospholipase D; Platelet Endothelial Cell Adhesion Molecule-1; Rats; Rats, Wistar; Receptors, Cannabinoid; Serotonin; TRPV Cation Channels

Dopamine and the endocannabinoids, anandamide and 2-arachidonoylglycerol, interact at several levels in the brain, with the involvement of both cannabinoid CB(1) receptors and transient receptor potential vanilloid type-1 (TRPV1) channels (which are alternative anandamide receptors). Using pharmacological, immunohistochemical and analytical approaches, we investigated the response of dopamine D(3) receptor null (D3R((-/-))) mice in models of epilepsy and anxiety, in relation to their brain endocannabinoid and endovanilloid tone. Compared to wild-type mice, D3R((-/-)) mice exhibited a delayed onset of clonic seizures, enhanced survival time, reduced mortality rate and more sensitivity to anticonvulsant effects of diazepam after intraperitoneal administration of picrotoxin (7 mg/kg), and a less anxious-like behaviour in the elevated plus maze test. D3R((-/-)) mice also exhibited different endocannabinoid and TRPV1, but not CB(1), levels in the hippocampus, nucleus accumbens, amygdala and striatum. Given the role played by CB(1) and TRPV1 in neuroprotection and anxiety, and based on data obtained here with pharmacological tools, we suggest that the alterations of endocannabinoid and endovanilloid tone found in D3R((-/-)) mice might account for part of their altered responses to excitotoxic and anxiogenic stimuli.

Topics: Analysis of Variance; Animals; Anti-Anxiety Agents; Anti-Inflammatory Agents, Non-Steroidal; Anxiety; Arachidonic Acids; Brain; Cannabinoid Receptor Modulators; Capsaicin; Diazepam; Disease Models, Animal; Dose-Response Relationship, Drug; Endocannabinoids; Epilepsy; GABA Antagonists; Male; Maze Learning; Mice; Mice, Inbred C57BL; Mice, Knockout; Picrotoxin; Piperidines; Pyrazoles; Reaction Time; Receptor, Cannabinoid, CB1; Receptors, Dopamine D3; Serotonin; TRPV Cation Channels

Knockout (KO) mice invalidated for the dopamine transporter (DAT) constitute a powerful animal model of neurobiological alterations associated with hyperdopaminergia relevant to schizophrenia and attention-deficit/hyperactivity disorder (ADHD).. Because of continuously increasing evidence for a neuromodulatory role of endocannabinoids in dopamine-related pathophysiological responses, we assessed endocannabinoid signaling in DAT KO mice and evaluated the ability of endocannabinoid ligands to normalize behavioral deficits, namely spontaneous hyperlocomotion in these mice.. In DAT KO mice, we found markedly reduced anandamide levels, specifically in striatum, the dopamine nerve terminal region. Furthermore, three distinct indirect endocannabinoid agonists, the selective anandamide reuptake inhibitors AM404 and VDM11 and the fatty acid amidohydrolase inhibitor AA5HT, attenuated spontaneous hyperlocomotion in DAT KO mice. The hypolocomotor effects of AM404, VDM11, and AA5HT were significantly attenuated by co-administration of the transient receptor potential vanilloid 1 (TRPV1) antagonist capsazepine but not the selective cannabinoid type 1 (CB1)receptor antagonist AM251. Interestingly, TRPV1 binding was increased in the striatum of DAT KO mice, while CB1 receptor binding was unaffected.. These data indicate a dysregulated striatal endocannabinoid neurotransmission associated with hyperdopaminergic state. Restoring endocannabinoid homeostasis in active synapses might constitute an alternative therapeutic strategy for disorders associated with hyperdopaminergia. In this process, TRPV1 receptors seem to play a key role and represent a novel promising pharmacological target.

Topics: Amidohydrolases; Animals; Arachidonic Acids; Attention Deficit Disorder with Hyperactivity; Cannabinoid Receptor Modulators; Capsaicin; Corpus Striatum; Disease Models, Animal; Dopamine; Dopamine Plasma Membrane Transport Proteins; Endocannabinoids; Mice; Mice, Knockout; Motor Activity; Polyunsaturated Alkamides; Receptor, Cannabinoid, CB1; Serotonin; Signal Transduction; TRPV Cation Channels

Direct stimulation of cannabinoid CB1 receptors exerts a protective function in animal models of inflammatory bowel diseases (IBDs). However, it is not known whether endocannabinoids are up-regulated during IBDs in animals or humans, nor whether pharmacological elevation of endocannabinoid levels can be exploited therapeutically in these disorders. In this study we addressed these questions. Colon inflammation was induced in mice and rats with 2,4-dinitrobenzene- and 2,4,6-trinitrobenzene sulfonic acids (DNBS and TNBS), respectively. DNBS-treated mice were treated chronically (for 3 or 7 days) with inhibitors of anandamide enzymatic hydrolysis (N-arachidonoyl-serotonin, AA-5-HT) or reuptake (VDM11), 10 or 5 mg/kg, s.c., or with 5-amino-salicilic acid (5-ASA, 1.4 mg/kg, i.r.). Endocannabinoids (anandamide and 2-arachidonoylglycerol, 2-AG) were quantified in mouse colon, or in rat colon mucosa and submucosa, and in bioptic samples from the colon of patients with untreated ulcerative colitis, by liquid chromatography-mass spectrometry. A strong elevation of anandamide, but not 2-AG, levels was found in the colon of DNBS-treated mice, in the colon submucosa of TNBS-treated rats, and in the biopsies of patients with ulcerative colitis. VDM-11 significantly elevated anandamide levels in the colon of DNBS-treated mice and concomitantly abolished inflammation, whereas AA-5-HT did not affect endocannabinoid levels and was significantly less efficacious at attenuating colitis. 5-ASA also increased anandamide levels and abolished colitis. Thus, anandamide is elevated in the inflamed colon of patients with ulcerative colitis, as well as in animal models of IBDs, to control inflammation, and elevation of its levels with inhibitors of its cellular reuptake might be used in the treatment of IBDs.

Topics: Adult; Aged; Amidohydrolases; Animals; Arachidonic Acids; Benzenesulfonates; Colitis; Colitis, Ulcerative; Colon; Disease Models, Animal; Drug Evaluation, Preclinical; Endocannabinoids; Female; Glycerides; Humans; Inflammatory Bowel Diseases; Intestinal Mucosa; Male; Mesalamine; Mice; Mice, Inbred C57BL; Middle Aged; Peroxidase; Polyunsaturated Alkamides; Rats; Rats, Wistar; Receptor, Cannabinoid, CB1; Serotonin; Specific Pathogen-Free Organisms; Trinitrobenzenesulfonic Acid