eCB Anandamide Research Articles

Sexually dimorphic effects of monoacylglycerol lipase inhibitor MJN110 on stress-related behaviour and drinking in Marchigian Sardinian alcohol-preferring rats.

The endocannabinoid (eCB) system plays an important homeostatic role in the regulation of stress circuits and has emerged as a therapeutic target to treat stress disorders and alcohol use disorder (AUD). Extensive research has elucidated a role for the eCB anandamide (AEA), but less is known about 2-arachidonoylglycerol (2-AG) mediated signalling. We pharmacologically enhanced eCB signalling by inhibiting the 2-AG metabolizing enzyme, monoacylglycerol lipase (MAGL), in male and female Marchigian Sardinian alcohol-preferring (msP) rats, a model of innate alcohol preference and stress hypersensitivity, and in control Wistar rats. We tested the acute effect of the selective MAGL inhibitor MJN110 in alleviating symptoms of alcohol drinking, anxiety, irritability and fear. A single systemic administration of MJN110 increased 2-AG levels in the central amygdala, prelimbic and infralimbic cortex but did not acutely alter alcohol drinking. MAGL inhibition reduced aggressive behaviours in female msPs, and increased defensive behaviours in male msPs, during the irritability test. Moreover, in the novelty-induced hypophagia test, MJN110 selectively enhanced palatable food consumption in females, mitigating stress-induced food suppression. Lastly, msP rats showed increased conditioned fear behaviour compared with Wistar rats, and MJN110 reduced context-associated conditioned fear responses, but not cue-probed fear expression, in male msPs. Acute inhibition of MAGL attenuated some stress-related responses in msP rats but not voluntary alcohol drinking. Our results provide new insights into the sex dimorphism documented in stress-induced responses. Sex-specific eCB-based approaches should be considered in the clinical development of therapeutics.

Structural basis for activation of CB1 by an endocannabinoid analog

Endocannabinoids (eCBs) are endogenous ligands of the cannabinoid receptor 1 (CB1), a G protein-coupled receptor that regulates a number of therapeutically relevant physiological responses. Hence, understanding the structural and functional consequences of eCB-CB1 interactions has important implications for designing effective drugs targeting this receptor. To characterize the molecular details of eCB interaction with CB1, we utilized AMG315, an analog of the eCB anandamide to determine the structure of the AMG315-bound CB1 signaling complex. Compared to previous structures, the ligand binding pocket shows some differences. Using docking, molecular dynamics simulations, and signaling assays we investigated the functional consequences of ligand interactions with the “toggle switch” residues F2003.36 and W3566.48. Further, we show that ligand-TM2 interactions drive changes to residues on the intracellular side of TM2 and are a determinant of efficacy in activating G protein. These intracellular TM2 rearrangements are unique to CB1 and are exploited by a CB1-specific allosteric modulator.

Sphingosine Kinases at the Intersection of Pro-Inflammatory LPS and Anti-Inflammatory Endocannabinoid Signaling in BV2 Mouse Microglia Cells.

Microglia, the resident immune cells of the central nervous system, play important roles in brain homeostasis as well as in neuroinflammation, neurodegeneration, neurovascular diseases, and traumatic brain injury. In this context, components of the endocannabinoid (eCB) system have been shown to shift microglia towards an anti-inflammatory activation state. Instead, much less is known about the functional role of the sphingosine kinase (SphK)/sphingosine-1-phosphate (S1P) system in microglia biology. In the present study, we addressed potential crosstalk of the eCB and the S1P systems in BV2 mouse microglia cells challenged with lipopolysaccharide (LPS). We show that URB597, the selective inhibitor of fatty acid amide hydrolase (FAAH)-the main degradative enzyme of the eCB anandamide-prevented LPS-induced production of tumor necrosis factor-α (TNFα) and interleukin-1β (IL-1β), and caused the accumulation of anandamide itself and eCB-like molecules such as oleic acid and cis-vaccenic acid ethanolamide, palmitoylethanolamide, and docosahexaenoyl ethanolamide. Furthermore, treatment with JWH133, a selective agonist of the eCB-binding cannabinoid 2 (CB2) receptor, mimicked the anti-inflammatory effects of URB597. Interestingly, LPS induced transcription of both SphK1 and SphK2, and the selective inhibitors of SphK1 (SLP7111228) and SphK2 (SLM6031434) strongly reduced LPS-induced TNFα and IL-1β production. Thus, the two SphKs were pro-inflammatory in BV2 cells in a non-redundant manner. Most importantly, the inhibition of FAAH by URB597, as well as the activation of CB2 by JWH133, prevented LPS-stimulated transcription of SphK1 and SphK2. These results present SphK1 and SphK2 at the intersection of pro-inflammatory LPS and anti-inflammatory eCB signaling, and suggest the further development of inhibitors of FAAH or SphKs for the treatment of neuroinflammatory diseases.

Endocannabinoid System Components of the Female Mouse Reproductive Tract Are Modulated during Reproductive Aging.

The endocannabinoid (eCB) system has gained ground as a key modulator of several female fertility-related processes, under physiological/pathological conditions. Nevertheless, its modulation during reproductive aging remains unclear. This study aimed to investigate the expression levels of the main receptors (cannabinoid receptor 1,CB1; cannabinoid receptor 2, CB2; G-protein coupled receptor, GPR55; and transient receptor potential vanilloid type 1 channel, TRPV1) and metabolic enzymes (N-acylphosphatidylethanolamine phospholipase D, NAPE-PLD; fatty acid amide hydrolase, FAAH; monoacylglycerol lipase, MAGL; and diacylglycerol lipase, DAGL-α and -β) of this system in the ovaries, oviducts, and uteri of mice at prepubertal, adult, late reproductive, and post-reproductive stages through quantitative ELISA and immunohistochemistry. The ELISA showed that among the receptors, TRPV1 had the highest expression and significantly increased during aging. Among the enzymes, NAPE-PLD, FAAH, and DAGL-β were the most expressed in these organs at all ages, and increased age-dependently. Immunohistochemistry revealed that, regardless of age, NAPE-PLD and FAAH were mainly found in the epithelial cells facing the lumen of the oviduct and uteri. Moreover, in ovaries, NAPE-PLD was predominant in the granulosa cells, while FAAH was sparse in the stromal compartment. Of note, the age-dependent increase in TRPV1 and DAGL-β could be indicative of increased inflammation, while that of NAPE-PLD and FAAH could suggest the need to tightly control the levels of the eCB anandamide at late reproductive age. These findings offer new insights into the role of the eCB system in female reproduction, with potential for therapeutic exploitation.

Coordinated Regulation of CB1 Cannabinoid Receptors and Anandamide Metabolism Stabilizes Network Activity during Homeostatic Downscaling.

Neurons express overlapping homeostatic mechanisms to regulate synaptic function and network properties in response to perturbations of neuronal activity. Endocannabinoids (eCBs) are bioactive lipids synthesized in the postsynaptic compartments to regulate synaptic transmission, plasticity, and neuronal excitability primarily through retrograde activation of presynaptic cannabinoid receptor type 1 (CB1). The eCB system is well situated to regulate neuronal network properties and coordinate presynaptic and postsynaptic activity. However, the role of the eCB system in homeostatic adaptations to neuronal hyperactivity is unknown. To address this issue, we used Western blotting and targeted lipidomics to measure adaptations in eCB system to bicuculline (BCC)-induced chronic hyperexcitation in mature cultured rat cortical neurons, and used multielectrode array (MEA) recording and live-cell imaging of glutamate dynamics to test the effects of pharmacological manipulations of eCB on network activities. We show that BCC-induced chronic hyperexcitation triggers homeostatic downscaling and a coordinated adaptation to enhance tonic eCB signaling. Hyperexcitation triggers first the downregulation of fatty acid amide hydrolase (FAAH), the lipase that degrades the eCB anandamide, then an accumulation of anandamide and related metabolites, and finally a delayed upregulation of surface and total CB1. Additionally, we show that BCC-induced downregulation of surface AMPA-type glutamate receptors (AMPARs) and upregulation of CB1 occur through independent mechanisms. Finally, we show that endocannabinoids support baseline network activities before and after downscaling and is engaged to suppress network activity during adaptation to hyperexcitation. We discuss the implications of our findings in the context of downscaling and homeostatic regulation of in vitro oscillatory network activities.

Tonic Endocannabinoid Levels Modulate Retinal Signaling.

The endocannabinoid (eCB) system is critically involved in the modulation of synaptic transmission in the central nervous system, playing an important role in the control of emotional responses, neurodevelopment and synaptic plasticity among other functions. The eCB system is also present in the retina, with studies indicating changes in function after application of cannabinoid receptor agonists, antagonists and in knockout models. Whether eCBs are tonically released in the retina and their physiological functions is, however, still unknown. We investigated the role of the eCB system in the modulation of response strength of retinal ganglion cells (RGCs) to light stimulation, their receptive field organization, contrast sensitivity and excitability properties by performing whole-cell patch-clamp recordings in mouse RGCs before and after bath application of URB597, an inhibitor of the enzyme that degrades the eCB anandamide. Our results show that URB597 application leads to a reduction in the strength of synaptic inputs onto RGCs but paradoxically increases RGC excitability. In addition, URB597 was shown to modulate receptive field organization and contrast sensitivity of RGCs. We conclude that tonically released eCBs modulate retinal signaling by acting on traditional cannabinoid receptors (CB1R/CB2R) as well as on non-cannabinoid receptor targets. Thus, a thorough understanding of the effects of drugs that alter the endogenous cannabinoid levels and of exogenous cannabinoids is necessary to fully comprehend the impact of their medical as well as recreational use on vision.

Endocannabinoids Modulate Production of Osteoclastogenic Factors by Stem Cells of the Apical Papilla In Vitro

IntroductionMany mediators are produced during pulp inflammation and necrosis, including endocannabinoids (ECbs), which might affect the function of stem cells of the apical papilla (SCAP), cells of paramount importance for root formation, and regenerative endodontic treatment. The aim of this study was to evaluate the production of osteoclastogenesis-related mediators by SCAP modulated by ECbs and lipopolysaccharide (LPS) in vitro. MethodsSCAP were cultured and treated with ECb anandamide (AEA), 2-arachidonoylglycerol, or N-arachidonoylaminophenol. All groups were incubated in the presence of a vehicle or LPS and the antagonist of transient receptor potential cation channel subfamily V member 1, capsazepine. After 24 hours, the culture medium supernatants were collected for further quantification of tumor necrosis factor alpha, CCL2, macrophage colony-stimulating factor, osteoprotegerin, and receptor activator of nuclear factor kappa B ligand. ResultsSmall amounts of tumor necrosis factor alpha and receptor activator of nuclear factor kappa B ligand were detected in SCAP supernatants, and none of the experimental conditions altered their production. A down-regulation in constitutive CCL2 production was observed in the AEA group compared with that in the LPS group. The production of macrophage colony-stimulating factor was significantly increased in all groups treated with AEA compared with the control and LPS-treated groups. Osteoprotegerin was significantly increased by AEA alone and by 2-arachidonoylglycerol and N-arachidonoylaminophenol in the presence of LPS and capsazepine. ConclusionsAEA modulates some of the osteoclastogenic factors produced by SCAP in a bone resorption protective fashion.

Measuring the Content of Endocannabinoid-Like Compounds in Biological Fluids: A Critical Overview of Sample Preparation Methodologies.

Different mass spectrometric techniques have been used over the past decade to quantify endocannabinoids (eCBs) and related lipids. Even with the level of molecular fingerprinting accuracy of an instrument like the most advanced triple quadrupole mass spectrometer, if one is not getting the most optimized sample to the detector in a way that this improved technology can be of use, then advancements can be stymied. Here, our focus is on review and discussion of sample preparation methodologies used to isolate the eCB anandamide and its close congeners N-acyl ethanolamines and structural congeners (i.e., lipo amino acids, lipoamines, N-acyl amides) in biological fluids. Most of our focus will be on the analysis of these lipids in plasma/serum, but we will also discuss how the same techniques can be used for the analysis of saliva and breast milk.

Simultaneous Assessment of Serum Levels and Pharmacologic Effects of Cannabinoids on Endocannabinoids and N-Acylethanolamines by Liquid Chromatography-Tandem Mass Spectrometry.

Introduction: The primary compounds of Cannabis sativa, delta-9-tetrahydrocannabinol (Δ9-THC) and cannabidiol (CBD), inflict a direct influence on the endocannabinoid system-a complex lipid signaling network with a central role in neurotransmission and control of inhibitory and excitatory synapses. These phytocannabinoids often interact with endogenously produced endocannabinoids (eCBs), as well as their structurally related N-acylethanolamines (NAEs), to drive neurobiological, nociceptive, and inflammatory responses. Identifying and quantifying changes in these lipid neuromodulators can be challenging owing to their low abundance in complex matrices. Materials and Methods: This article describes a robust liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the extraction and quantification of the eCBs anandamide and 2-arachidonoylglycerol, along with their congener NAEs oleoylethanolamine and palmitoylethanolamine, and phytocannabinoids CBD, Δ9-THC, and 11-Nor-9-carboxy-Δ9-tetrahydrocannabinol, a major metabolite of Δ9-THC. Our method was applied to explore pharmacokinetic and pharmacodynamic effects from intraperitoneal injections of Δ9-THC and CBD on circulating levels of eCBs and NAEs in rodent serum. Results: Detection limits ranged from low nanomolar to picomolar in concentration for eCBs (0.012-0.24 pmol/mL), NAEs (0.059 pmol/mL), and phytocannabinoids (0.24-0.73 pmol/mL). Our method displayed good linearity for calibration curves of all analytes (R2>0.99) as well as acceptable accuracy and precision, with quality controls not deviating >15% from their nominal value. Our LC-MS/MS method reliably identified changes to these endogenous lipid mediators that followed a causal relationship, which was dependent on both the type of phytocannabinoid administered and its pharmaceutical preparation. Conclusion: We present a rapid and reliable method for the simultaneous quantification of phytocannabinoids, eCBs, and NAEs in serum using LC-MS/MS. The accuracy and sensitivity of our assay infer it can routinely monitor endogenous levels of these lipid neuromodulators in serum and their response to external stimuli, including cannabimimetic agents.

Activation of TRPV1 by Capsaicin or Heat Drives Changes in 2-Acyl Glycerols and N-Acyl Ethanolamines in a Time, Dose, and Temperature Dependent Manner.

Endocannabinoids (eCBs) and transient receptor potential (TRP) channels are associated with thermoregulation; however, there are many gaps in the understanding of how these signaling systems work together in responding to changes in temperature. TRPV1, a calcium-permeable ion channel, is activated by capsaicin, elevated temperature, the eCB Anandamide, and over 15 additional endogenous lipids. There is also evidence for signaling crosstalk between TRPV1 and the eCB receptor, CB1. We recently found that activation of TRPV1-HEK cells by capsaicin increases the production of the eCB, 2-arachidonoyl glycerol (2-AG), suggesting a molecular link between these receptors. Here, we tested the hypothesis that TRPV1 activation by capsaicin drives regulation of a wider-range of lipid signaling molecules and is time and dose-dependent. We also tested the hypothesis that changes in temperature that drive changes in calcium mobilization in TRPV1-HEK will likewise drive similar changes in lipid signaling molecule regulation. Lipid analysis was conducted by partial purification of methanolic extracts on C18 solid phase extraction columns followed by HPLC/MS/MS. Capsaicin increased the release of 2-acyl glycerols (2-AG, 2-linoleoyl glycerol, 2-oleoyl glycerol), in a concentration- and time-dependent manner, whereas levels of N-acyl ethanolamines (NAEs), including Anandamide, were significantly decreased. Analogous changes in 2-acyl glycerols and NAEs were measured upon ramping the temperature from 37 to 45°C. In contrast, opposite effects were measured when analyzing lipids after they were maintained at 27°C and then quickly ramped to 37°C, wherein 2-acyl glycerol levels decreased and NAEs increased. These results provide further evidence that the eCB system and TRPV1 have integrated signaling functions that are associated with the molecular response to temperature variation.

Exercise-induced euphoria and anxiolysis do not depend on endogenous opioids in humans

A runner's high describes a sense of well-being during endurance exercise characterized by euphoria and anxiolysis. It has been a widespread belief that the release of endogenous opioids, such as endorphins, underlie a runner's high. However, exercise leads to the release of two classes of rewarding molecules, endocannabinoids (eCBs) and opioids. In mice, we have shown that core features of a runner's high depend on cannabinoid receptors but not opioid receptors. In the present study, we aimed to corroborate in humans that endorphins do not play a significant role in the underlying mechanism of a runner's high. Thus, we investigated whether the development of two core features of a runner's high, euphoria and reduced anxiety levels, depend on opioid signaling by using the opioid receptor antagonist naltrexone (NAL) in a double-blind, randomized, placebo (PLA)-controlled experiment. Participants (N = 63) exhibited increased euphoria and decreased anxiety after 45 min of running (RUN) on a treadmill in a moderate-intensity range compared to walking (WALK). RUN led to higher plasma levels of the eCBs anandamide (AEA) and 2-arachidonoglycerol (2-AG). Opioid blockade did not prevent the development of euphoria and reduced anxiety as well as elevation of eCB levels following exercise. Moreover, the fraction of participants reporting a subjective runner's high was comparable in the NAL and PLA-treated group. Therefore, this study indicates that the development of a runner's high does not depend on opioid signaling in humans, but makes eCBs strong candidates in humans, as previously shown in mice.

The endocannabinoid system - current implications for drug development.

In this review, the state of the art for compounds affecting the endocannabinoid (eCB) system is described with a focus on the treatment of pain. Amongst directly acting CB receptor ligands, clinical experience with ∆9‐tetrahydracannabinol and medical cannabis in chronic non‐cancer pain indicates that there are differences between the benefits perceived by patients and the at best modest effect seen in meta‐analyses of randomized controlled trials. The reason for this difference is not known but may involve differences in the type of patients that are recruited, the study conditions that are chosen and the degree to which biases such as reporting bias are operative. Other directly acting CB receptor ligands such as biased agonists and allosteric receptor modulators have not yet reached the clinic. Amongst indirectly acting compounds targeting the enzymes responsible for the synthesis and catabolism of the eCBs anandamide and 2‐arachidonoylglycerol, fatty acid amide hydrolase (FAAH) inhibitors have been investigated clinically but were per se not useful for the treatment of pain, although they may be useful for the treatment of post‐traumatic stress disorder and cannabis use disorder. Dual‐acting compounds targeting this enzyme and other targets such as cyclooxygenase‐2 or transient potential vanilloid receptor 1 may be a way forward for the treatment of pain.

Comorbid anxiety-like behavior in a rat model of colitis is mediated by an upregulation of corticolimbic fatty acid amide hydrolase

Peripheral inflammatory conditions, including those localized to the gastrointestinal tract, are highly comorbid with psychiatric disorders such as anxiety and depression. These behavioral symptoms are poorly managed by conventional treatments for inflammatory diseases and contribute to quality of life impairments. Peripheral inflammation is associated with sustained elevations in circulating glucocorticoid hormones, which can modulate central processes, including those involved in the regulation of emotional behavior. The endocannabinoid (eCB) system is exquisitely sensitive to these hormonal changes and is a significant regulator of emotional behavior. The impact of peripheral inflammation on central eCB function, and whether this is related to the development of these behavioral comorbidities remains to be determined. To examine this, we employed the trinitrobenzene sulfonic acid-induced model of colonic inflammation (colitis) in adult, male, Sprague Dawley rats to produce sustained peripheral inflammation. Colitis produced increases in behavioral measures of anxiety and elevations in circulating corticosterone. These alterations were accompanied by elevated hydrolytic activity of the enzyme fatty acid amide hydrolase (FAAH), which hydrolyzes the eCB anandamide (AEA), throughout multiple corticolimbic brain regions. This elevation of FAAH activity was associated with broad reductions in the content of AEA, whose decline was driven by central corticotropin releasing factor type 1 receptor signaling. Colitis-induced anxiety was reversed following acute central inhibition of FAAH, suggesting that the reductions in AEA produced by colitis contributed to the generation of anxiety. These data provide a novel perspective for the pharmacological management of psychiatric comorbidities of chronic inflammatory conditions through modulation of eCB signaling.

The endocannabinoidome as a substrate for noneuphoric phytocannabinoid action and gut microbiome dysfunction in neuropsychiatric disorders .

The endocannabinoid (eCB) system encompasses the eCBs anandamide and 2-arachidonoylglycerol, their anabolic/catabolic enzymes, and the cannabinoid CB1 and CB2 receptors. Its expansion to include several eCB-like lipid mediators, their metabolic enzymes, and their molecular targets, forms the endocannabinoidome (eCBome). This complex signaling system is deeply involved in the onset, progress, and symptoms of major neuropsychiatric disorders and provides a substrate for future therapeutic drugs against these diseases. Such drugs may include not only THC, the major psychotropic component of cannabis, but also other, noneuphoric plant cannabinoids. These compounds, unlike THC, possess a wide therapeutic window, possibly due to their capability of hitting several eCBome and non-eCBome receptors. This is particularly true for cannabidiol, which is one of the most studied cannabinoids and shows promise for the treatment of a wide range of mental and mood disorders. The eCBome plays a role also in the microbiota-gut-brain axis, which is emerging as an important actor in the control of affective and cognitive functions and in their pathological alterations.

HYPNOTIZABILITY-RELATED FAAH C385A POLYMORPHISM: POSSIBLE ENDOCANNABINOID CONTRIBUTION TO SUGGESTION-INDUCED ANALGESIA

ABSTRACT Fatty acid amide hydrolase (FAAH) degrades the endogenous endocannabinoid (eCB) anandamide and might be involved in the response to suggestions of analgesia in subjects with high hypnotizability scores (highs). Since the A allele of the FAAH C385A polymorphism (rs324420) is associated with lower FAAH activity, it was studied in 21 highs, 66 low hypnotizable individuals (lows), and 172 individuals not selected for hypnotizability (controls) representing the general population. No significant difference was observed among groups, but the A allele frequency showed a significant trend to increase from lows to controls and from controls to highs. Since eCB small differences can be amplified by eCB interactions with other neurotransmitters, a contribution of the FAAH polymorphism to the highs’ analgesia should not be excluded.

Altered circulating endocannabinoids in anorexia nervosa during acute and weight-restored phases: A pilot study.

Anorexia nervosa (AN) is an eating disorder characterized by a low food intake and often exceeding exercise, leading to a particularly low body × weight proportion. Patients with AN usually report less hunger than healthy controls. Endogenous endocannabinoids (eCBs), specifically the anandamide, have been associated to hunger, as a meal initiator, but research regarding AN and eCB and inconclusive. In this pilot study, we investigated plasma levels of eCB in inpatients with AN during fasting and after eating, both during the acute AN phase and after weight recovery. After an 8-hr fasting period, blood sample was collected from all participants. After that, participants were given a muffin test meal. Blood samples for the investigation of endogenous eCBs anandamide (N-arachidonoylethanolamide [AEA]) and 2-arachidonoylglycerol (2-AG) were then collected after 120 and 240 min. Participants were only allowed to eat and drink what was offered them during the research. AN reported less hunger than controls during fasting and at the end of the experiment. Also, plasma levels of AEA were significantly smaller in AN in comparison with controls in all time points. No significant difference was found for 2-AG plasma levels. After recovery, no significant difference was found for eCB levels. These findings could be interpreted as an AEA deregulation in AN before and after food intake, which persists after weight recovery. These findings may have implications to the pharmacological treatment of AN and to relapse occurring in the disorder.

Endocannabinoids Interact With the Dopaminergic System to Increase Sexual Motivation: Lessons From the Sexual Satiety Phenomenon.

In male rats, copulation to satiety induces a long-lasting sexual inhibitory state, considered to rely on a decreased sexual motivation. Dopaminergic transmission at the mesolimbic system plays a central role in the regulation of male sexual motivation. Endocannabinoids (eCBs) modulate the activity of the mesolimbic system and both dopamine (DA) and cannabinoid receptor activation reverses the sexual inhibition that characterizes sexually satiated rats. The eCB anandamide reverses sexual satiety when systemically administered or infused into the ventral tegmental area (VTA), the region where the activity of mesolimbic dopaminergic neurons is regulated. Thus, it could be thought that sexual motivation is diminished during the long-lasting sexual inhibition of sexually satiated rats and that eCBs reverse that inhibition through the modulation of the dopaminergic system. To test this hypothesis, we assessed the motivational state of sexually satiated male rats and determined if 2-arachidonoylglycerol (2-AG), the most abundant eCB and a full cannabinoid receptor agonist, also reversed the sexual inhibitory state. To establish the possible interaction between 2-AG and anandamide with the dopaminergic system for the reversal of sexual satiety, we analyzed the effects of the co-administration of each eCB and DA receptor agonists or antagonists. Results showed that 24-h after copulation to satiety, when the sexual inhibition is well established, the males’ sexual motivation is diminished as measured in the sexual incentive motivation test. 2-AG, similarly to anandamide, reverses sexual satiety through the activation of CB1 receptors and both eCBs interact with the dopaminergic system to reverse the sexual inhibitory state. 2-AG effects are mediated by the modulation of the D2-like DA receptor family, whereas anandamide’s effects are clearly mediated by the modulation of the D1-like DA receptor family and the activation of D2-like DA receptors. Present results evidence that a reduced sexual motivation underlies the sexual inhibitory state of sexually satiated rats and support the notion that eCBs reverse sexual satiety by modulating dopaminergic transmission, presumably at the mesolimbic system. Anandamide and 2-AG have a different interaction with D1-like and D2-like DA receptor families. Altogether present data endorse the association of the eCB system with the regulation of the motivational tone at the mesolimbic system.

S31. Beneficial Effects of FAAH Inhibition on Fear- and Stress-Related Behaviors in Healthy Humans

The endocannabinoid (eCB) system is proposed to act as a “stress buffer” and thus represents a novel therapeutic target for stress-related psychiatric disorders. Preclinical data suggests that inhibition of fatty acid amide hydrolase (FAAH), the main degradative enzyme of the eCB anandamide (AEA), facilitates fear extinction and protects against the anxiogenic effects of stress. However, no studies have yet assessed whether pharmacological inhibition of FAAH would produce similar effects in humans.

Hemisphere-dependent endocannabinoid system activity in prefrontal cortex and hippocampus of the Flinders Sensitive Line rodent model of depression

Altered endocannabinoid (eCB) signalling is suggested as an important contributor to the pathophysiology of depression. To further elucidate this, we conducted a study using a genetic rat model of depression, the Flinders Sensitive Line (FSL), and their controls, the Flinders Resistant Line (FRL) rats. Plasma, right and left prefrontal cortex, and hippocampus were isolated from FSL and FRL rats. We analyzed each region for the eCB anandamide (AEA) and 2-arachidonoylglycerol (2-AG) levels by liquid chromatography/multiple reaction monitoring (LC/MRM), mRNA and protein levels of the cannabinoid type 1 receptor (CB1R), fatty acid amide hydrolase (FAAH) and monoacyl glycerol lipase (MAGL) by real time qPCR and Western blotting. Content of 2-AG was lower in the left side of the hippocampus and prefrontal cortex in FSL rats compared to FRL rats. Inversely, levels of AEA were higher in right hippocampus than in left hippocampus. In plasma, AEA levels were increased and 2-AG decreased. Cannabinoid receptor 1 (Cnr1), Faah and Magl mRNA levels were prominently decreased in right prefrontal cortex of FSL rats as compared to FRL rats. Protein expression of CB1R and FAAH were decreased in left hippocampus. In summary, our data suggest a decreased eCB signalling in the FSL rats, which could contribute to the depressive-like behaviour. Interestingly, the altered eCB system activity appear to be hemisphere-specific in the limbic regions. Our study support the existing literature and showed altered eCB system activity in this particular animal model of depression.

P.2.007 - Endocannabinoid interactions in the regulation of acute responses to trauma and formation of traumatic memories

Endocannabinoid (eCB) signaling is a major contributor in the regulation of behavioral responses to traumas [1] and acquisition of traumatic memories [2]. ECBs anandamide (AEA) and 2-arachidonoilglycerol (2-AG) were reported to differentially affect particular aspects of behaviour [3] but their specific role in the regulation of responses to traumatic events is yet to be clarified. Furthermore, AEA and 2-AG signaling were recently shown to interact [4] which can possibly have functional relevance in their behavioral effects as well. Here we aimed to describe the specific contribution of eCBs and their interactions in the regulation of acute responses to a traumatic event and the acquisition of traumatic memories. We employed electric footshocks to traumatize Wistar rats in which eCB signaling was enhanced by the specific pharmacological blockade of the degrading enzyme of AEA by URB597 and 2-AG by JZL184. Treatments were delivered either systemically (URB597: 0.3 mg/kg; JZL184: 16 mg/kg) or locally to brain areas relevant in the regulation of traumatic memory, the ventral hippocampus (vHC), prelimbic cortex (PrL) and basolateral amygdala (BLA) (URB597: 1 ng/0.5 ul; JZL184: 1 ug/0.5 ul). To elucidate whether the effects of enhanced eCB signaling are modulated via cannabinoid receptor type-1 (CB1R), CB1R antagonist AM251 (1 mg/kg) was simultaneously administered with URB597 and JZL184. Systemic administration of JZL148 dampened acute fear responses to footshocks measured by freezing behavior, which was blocked by parallel treatment with URB597 (one-way ANOVA: F(4,46)=13.59; p