Blockade Of Fatty Acid Amide Hydrolase Research Articles

Blockade of 2-Arachidonoylglycerol Hydrolysis by Selective Monoacylglycerol Lipase Inhibitor 4-Nitrophenyl 4-(Dibenzo[d][1,3]dioxol-5-yl(hydroxy)methyl)piperidine-1-carboxylate (JZL184) Enhances Retrograde Endocannabinoid Signaling

Endocannabinoid (eCB) signaling mediates depolarization-induced suppression of excitation (DSE) and inhibition (DSI), two prominent forms of retrograde synaptic depression. N-Arachidonoylethanolamine (AEA) and 2-arachidonoylglycerol (2-AG), two known eCBs, are degraded by fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL), respectively. Selective blockade of FAAH and MAGL is critical for determining the roles of the eCBs in DSE/DSI and understanding how their action is regulated. 4-Nitrophenyl 4-(dibenzo[d][1,3]dioxol-5-yl(hydroxy)methyl)piperidine-1-carboxylate (JZL184) is a recently developed, highly selective, and potent MAGL inhibitor that increases 2-AG but not AEA concentrations in mouse brain. Here, we report that JZL184 prolongs DSE in Purkinje neurons in cerebellar slices and DSI in CA1 pyramidal neurons in hippocampal slices. The effect of JZL184 on DSE/DSI is mimicked by the nonselective MAGL inhibitor methyl arachidonyl fluorophosphonate. In contrast, neither the selective FAAH inhibitor cyclohexylcarbamic acid 3'-carbomoylbiphenyl-3-yl ester (URB597) nor FAAH knockout has a significant effect on DSE/DSI. JZL184 produces greater enhancement of DSE/DSI in mouse neurons than that in rat neurons. The latter finding is consistent with biochemical studies showing that JZL184 is more potent in inhibiting mouse MAGL than rat MAGL. These results indicate that the degradation of 2-AG by MAGL is the rate-limiting step that determines the time course of DSE/DSI and that JZL184 is a useful tool for the study of 2-AG-mediated signaling.

Fatty acid amide hydrolase (FAAH) knockout mice exhibit enhanced acquisition of an aversive, but not of an appetitive, Barnes maze task

It is well established that genetic deletion or pharmacological inhibition of the CB 1 receptor disrupts extinction learning in aversive conditioning tasks, but not in appetitive tasks. Consistent with these findings is that genetic deletion or pharmacological inhibition of fatty acid amide hydrolase (FAAH), the primary catabolic enzyme of the endogenous cannabinoid anandamide (AEA), accelerates acquisition as well as extinction in aversive conditioning tasks. However, it is unknown whether FAAH blockade will affect acquisition in an appetitive conditioning task. Therefore, in the present study, we assessed FAAH (−/−) and (+/+) mice in appetitive and aversive Barnes maze conditioning procedures. Here we report that FAAH (−/−) mice displayed accelerated acquisition rates in an aversively-motivated, but not in the appetitively-motivated, Barnes maze task. The CB 1 receptor antagonist, rimonabant attenuated enhanced acquisition in the aversive procedure, consistent with the idea that elevated AEA levels mediate this apparent nootropic effect. These findings support the hypothesis that stimulation of the endocannabinoid system enhances learned behavior in aversive, but not appetitive, conditioning paradigms.

Targeting Fatty Acid Amide Hydrolase (FAAH) to Treat Pain and Inflammation

The endogenous cannabinoid N-arachidonoyl ethanolamine (anandamide; AEA) produces most of its pharmacological effects by binding and activating CB(1) and CB(2) cannabinoid receptors within the CNS and periphery. However, the actions of AEA are short lived because of its rapid catabolism by fatty acid amide hydrolase (FAAH). Indeed, FAAH knockout mice as well as animals treated with FAAH inhibitors are severely impaired in their ability to hydrolyze AEA as well as a variety of noncannabinoid lipid signaling molecules and consequently possess greatly elevated levels of these endogenous ligands. In this mini review, we describe recent research that has investigated the functional consequences of inhibiting this enzyme in a wide range of animal models of inflammatory and neuropathic pain states. FAAH-compromised animals reliably display antinociceptive and anti-inflammatory phenotypes with a similar efficacy as direct-acting cannabinoid receptor agonists, such as Delta(9)-tetrahydrocannabinol (THC), the primary psychoactive constituent of Cannabis sativa. Importantly, FAAH blockade does not elicit any apparent psychomimetic effects associated with THC or produce reinforcing effects that are predictive of human drug abuse. The beneficial effects caused by FAAH blockade in these models are predominantly mediated through the activation of CB(1) and/or CB(2) receptors, though noncannabinoid mechanisms of actions can also play contributory or even primary roles. Collectively, the current body of scientific literature suggests that activating the endogenous cannabinoid system by targeting FAAH is a promising strategy to treat pain and inflammation but lacks untoward side effects typically associated with Cannabis sativa.

Inhibiting parabrachial fatty acid amide hydrolase activity selectively increases the intake of palatable food via cannabinoid CB1 receptors

These studies investigated feeding responses to indirect activation of parabrachial cannabinoid CB1 receptors. Arachidonoyl serotonin (AA5HT), an inhibitor of the endocannabinoid degradative enzyme, fatty acid amide hydrolase (FAAH), was infused into the parabrachial nucleus of male Sprague-Dawley rats, and intakes of high-fat/sucrose pellets and standard rodent chow were subsequently evaluated under various feeding schedules. FAAH blockade stimulated the intake of high-fat/sucrose pellets that were presented daily for 4 h during the light period, with compensatory decreases in the consumption of standard chow during the ensuing 20 h. These diet-selective changes were repeated on the next day, indicating a shift in feeding toward the more palatable diet that lasted for 48 h after a single infusion. The cannabinoid CB1 receptor antagonist, AM251, blocked the orexigenic actions of AA5HT, implicating CB1 receptors in mediating the feeding responses to FAAH inactivation. When the feeding schedule was reversed, AA5HT produced nominal increases in the consumption of standard chow for the 4-h access period, but substantial increases in the intake of high-fat/sucrose during the following 20-h interval. When presented with only high-fat/sucrose diet for 24 h, AA5HT increased 24-h food intake. In contrast, when given 24-h access only to standard chow, AA5HT failed to affect intake. Therefore, indirectly activating parabrachial CB1 receptors by blocking the degradation of native ligands selectively stimulates the intake of palatable food, with differential actions on total energy intake depending upon the feeding schedule. Our results support a role for parabrachial cannabinoid mechanisms in providing physiological regulation to neural substrates modulating feeding, energy balance, and behavioral responses for natural reward.

Targeting FAAH and COX to Treat Visceral Pain

Genetic deletion or pharmacological inhibition of fatty acid amide hydrolase (FAAH), the primary enzyme responsible for catabolism of the endogenous cannabinoid anandamide, leads to a CB1 receptor mediated decrease in nociceptive behavior in a variety of inflammatory and neuropathic pain models. However, there are presently no published reports that have evaluated whether FAAH blockade reduces nociception in visceral pain, though direct‐acting CB1 receptor agonists as well as cyclooxygenase (COX) inhibitors are highly effective in visceral pain assays. Therefore, in the present study we evaluated whether FAAH blockade will block nociceptive behavior in the mouse acetic acid writhing model. Additionally, we examined whether co‐administration of a FAAH inhibitor and COX inhibitor would synergize to block visceral pain. FAAH (‐/‐) mice as well as wild type mice treated with FAAH inhibitors (i.e., URB597 and OL‐135) or the non‐selective COX inhibitor diclofenac displayed a decrease in acetic acid‐induced writhing. URB597 (1–10 mg/kg; ED50 = 2.1 mg/kg) and diclofenac (3–30 mg/kg; ED50 = 9.8 mg/kg) produced dose‐dependent antinociception in the writhing test. The CB1 receptor antagonist rimonabant blocked the analgesic effects in the FAAH‐compromised mice, but failed to attenuate the actions of diclofenac. Interactions between URB597 and diclofenac were analyzed using an isobolographic approach. Combined administration of URB59597 and diclofenac in ratios of 1:1 [ED50s = 0.24 and 1.18 mg/kg], 1:3 [ED50s = 0.28 and 4.30 mg/kg], or 3:1 yielded [ED50s = 0.44 and 0.72 mg/kg] yielded synergistic interactions. Taken together, the results of the present study suggest that FAAH represents a promising target to treat visceral pain and combination of FAAH inhibitors and NSAIDs may have greater therapeutic utility than either class of drug given alone.

Evaluation of fatty acid amides in the carrageenan-induced paw edema model

While it has long been recognized that Δ9-tetrahydrocannabinol (THC), the primary psychoactive constituent of cannabis, and other cannabinoid receptor agonists possess anti-inflammatory properties, their well known CNS effects have dampened enthusiasm for therapeutic development. On the other hand, genetic deletion of fatty acid amide hydrolase (FAAH), the enzyme responsible for degradation of fatty acid amides, including endogenous cannabinoid N-arachidonoyl ethanolamine (anandamide; AEA), N-palmitoyl ethanolamine (PEA), N-oleoyl ethanolamine (OEA), and oleamide, also elicits anti-edema, but does not produce any apparent cannabinoid effects. The purpose of the present study was to investigate whether exogenous administration of FAAs would augment the anti-inflammatory phenotype of FAAH (−/−) mice in the carrageenan model. Thus, we evaluated the effects of the FAAs AEA, PEA, OEA, and oleamide in wild-type and FAAH (−/−) mice. For comparison, we evaluated the anti-edema effects of THC, dexamethasone (DEX), a synthetic glucocorticoid, diclofenac (DIC), a nonselective cyclooxygenase (COX) inhibitor, in both genotypes. A final study determined if tolerance to the anti-edema effects of PEA occurs after repeated dosing. PEA, THC, DEX, DIC elicited significant decreases in carrageenan-induced paw edema in wild-type mice. In contrast OEA produced a less reliable anti-edema effect than these other drugs, and AEA and oleamide failed to produce any significant decreases in paw edema. Moreover, none of the agents evaluated augmented the anti-edema phenotype of FAAH (−/−) mice, suggesting that maximal anti-edema effects had already been established. PEA was the most effective FAA in preventing paw edema and its effects did not undergo tolerance. While the present findings do not support a role for AEA in preventing carrageenan-induced edema, PEA administration and FAAH blockade elicited anti-edema effects of an equivalent magnitude as produced by THC, DEX, and DIC in this assay.

Role of Endocannabinoids in Alcohol Consumption and Intoxication: Studies of Mice Lacking Fatty Acid Amide Hydrolase

Endocannabinoid signaling plays the important role in regulation of ethanol intake. Fatty acid amide hydrolase (FAAH) is a key membrane protein for metabolism of endocannabinoids, including anandamide, and blockade of FAAH increases the level of anandamide in the brain. To determine if FAAH regulates ethanol consumption, we studied mutant mice with deletion of the FAAH gene. Null mutant mice showed higher preference for alcohol and voluntarily consumed more alcohol than wild-type littermates. There was no significant difference in consumption of sweet or bitter solutions. To determine the specificity of FAAH for ethanol intake, we studied additional ethanol-related behaviors. There were no differences between null mutant and wild-type mice in severity of ethanol-induced acute withdrawal, conditioned taste aversion to alcohol, conditioned place preference, or sensitivity to hypnotic effect of ethanol. However, null mutant mice showed shorter duration of loss of righting reflex induced by low doses of ethanol (3.2 and 3.4 g/kg) and faster recovery from motor incoordination induced by ethanol. All three behavioral phenotypes (increased preference for ethanol, decreased sensitivity to ethanol-induced sedation, and faster recovery from ethanol-induced motor incoordination) seen in mutant mice were reproduced in wild-type mice by injection of a specific inhibitor of FAAH activity--URB597. These data suggest that increased endocannabinoid signaling increased ethanol consumption owing to decreased acute ethanol intoxication.

Anandamide administration alone and after inhibition of fatty acid amide hydrolase (FAAH) increases dopamine levels in the nucleus accumbens shell in rats

Although endogenous cannabinoid systems have been implicated in the modulation of the rewarding effects of abused drugs and food, little is known about the direct effects of endogenous ligands for cannabinoid receptors on brain reward processes. Here we show for the first time that the intravenous administration of anandamide, an endogenous ligand for cannabinoid receptors, and its longer-lasting synthetic analog methanandamide, increase the extracellular dopamine levels in the nucleus accumbens shell of awake, freely moving rats, an effect characteristic of most drugs abused by humans. Anandamide produced two distinctly different effects on dopamine levels: (1) a rapid, transient increase that was blocked by the cannabinoid CB1 receptor antagonist rimonabant, but not by the vanilloid VR1 receptor antagonist capsazepine, and was magnified and prolonged by the fatty acid amide hydrolase (FAAH) enzyme inhibitor, URB597; (2) a smaller delayed and long-lasting increase, not sensitive to CB1, VR1 or FAAH blockade. Both effects were blocked by infusing either tetrodotoxin (TTX, 1 microm) or calcium-free Ringer's solution through the microdialysis probe, demonstrating that they were dependent on the physiologic activation of dopaminergic neurotransmission. Thus, these results indicate that anandamide, through the activation of the mesolimbic dopaminergic system, participates in the signaling of brain reward processes.

Fatty Acid Amide Hydrolase Controls Mouse Intestinal Motility In Vivo

Fatty acid amide hydrolase (FAAH) catalyzes the hydrolysis both of the endocannabinoids (which are known to inhibit intestinal motility) and other bioactive amides (palmitoylethanolamide, oleamide, and oleoylethanolamide), which might affect intestinal motility. The physiologic role of FAAH in the gut is largely unexplored. In the present study, we evaluated the possible role of FAAH in regulating intestinal motility in mice in vivo. Motility was measured by evaluating the distribution of a fluorescent marker along the small intestine; FAAH messenger RNA (mRNA) levels were analyzed by reverse-transcription polymerase chain reaction (RT-PCR); endocannabinoid levels were measured by isotope-dilution, liquid chromatography, mass spectrometry. Motility was inhibited by N-arachidonoylserotonin (AA-5-HT) and palmitoylisopropylamide, 2 selective FAAH inhibitors, as well as by the FAAH substrates palmitoylethanolamide, oleamide, and oleoylethanolamide. The effect of AA-5-HT was reduced by the CB1 receptor antagonist rimonabant and by CB1 deficiency in mice but not by the vanilloid receptor antagonist 5'-iodoresiniferatoxin. In FAAH-deficient mice, pharmacologic blockade of FAAH did not affect intestinal motility. FAAH mRNA was detected in different regions of the intestinal tract. We conclude that FAAH is a physiologic regulator of intestinal motility and a potential target for the development of drugs capable of reducing intestinal motility.

Differential effects of the sleep-inducing lipid oleamide and cannabinoids on the induction of long-term potentiation in the CA1 neurons of the rat hippocampus in vitro

Cannabinoids have been shown to impair cognition in vivo and block long-term potentiation (LTP), a candidate experimental model of learning and memory in vitro, via cannabinoid receptor (CB1) activation. cis-Oleamide (cOA) is an endogenous sleep-inducing lipid with putative cannabinomimetic properties. We hypothesise that cOA is cannabinomimetic and perform a comparative study with synthetic and endogenous cannabinoids on their effects on synaptic conditioning via two different patterns of stimulation in the hippocampal slice. CB1 agonists, R(+)-WIN55212-2 and anandamide, but not cOA blocked high frequency stimulation (HFS)-LTP. R(+)-WIN55212-2 and cOA (stereoselectively) attenuated responses to θ-burst-LTP, while anandamide did not. The anandamide transport inhibitor, AM404, attenuated HFS-LTP, an effect reversed by the CB1 receptor antagonist SR141716A but not mimicked by the vanilloid receptor agonist capsaicin. TFNO, an inhibitor of fatty acid amide hydrolase (FAAH), the enzyme responsible for degrading anandamide, failed to block HFS-LTP alone or in combination with cOA. On the contrary, this combination was as effective as cOA on its own in attenuating θ-burst-LTP. cOA effects on θ-burst-LTP were prevented in the presence of the GABA A receptor blocker picrotoxin, but not by pretreatment with SR141716A. These findings suggest that cOA neither directly activates CB1 receptors nor acts via the proposed “entourage” effect [Nature 389 (1997) 25] to increase titres of anandamide through FAAH inhibition. The selective effects of cOA on θ-burst-conditioning may reflect modulation of GABAergic transmission. Anandamide uptake inhibition, but not blockade of FAAH, effectively increases synaptic concentrations of endocannabinoids.