This study aimed to evaluate whether salivary endocannabinoid (EC) and N-acylethanolamine (NAE) concentrations upon mastication of a semisolid food were involved in the sensory perception of fat taste, food liking and appetite in humans. A fat-enriched (FEP) and a low-fat control (CP) pudding were developed and used in a randomized cross-over study with 19 healthy volunteers. The study protocol combined a Modified Sham-Feeding (MSF) with a multiple-spoon Temporal Dominance of Sensations method. Subjects masticated and expectorated 10 spoons of the pudding and selected the dominant sensations among a list of attributes. Saliva samples, appetite and food liking scores were collected at baseline, immediately after the MSF of the pudding and every 5 min until 20 min after MSF. Salivary concentrations of all monitored ECs and NAEs increased during pudding mastication compared to baseline (except for palmitoylethanolamide with FEP). The raise was lower with FEP than with CP for all compounds except for 2-arachidonoylglycerol whose increase was higher than the other compounds and independent of pudding type. Salivary N-arachidonoylethanolamine, linoleoylethanolamide and palmitoylethanolamide were significantly lower at 10 and 20 min after MSF of FEP than CP. Fatty taste at the 2nd spoon and creaminess at the 5th spoon were perceived as dominant with FEP whereas only wateriness was dominant with CP at the 2nd spoon. No difference between puddings for individual appetite or food liking over the 20 min of the protocol was recorded. During mastication of a semisolid fat-enriched food, the fatty taste and the creaminess were perceived as dominant. Salivary ECs and NAEs were not associated with the individual perception of fatty taste, pudding liking and appetite sensations.

Endocannabinoids (ECs) represent a class of endogenous, small molecules that bind and activate the G-protein coupled EC receptors. They are involved in a variety of fundamental biological processes and are associated with many disease states. Endocannabinoids are often present in complex matrices and at low concentrations, complicating their measurement. Here we describe a highly sensitive method for the quantitation of the following ECs in serum: N-arachidonoylethanolamine (anandamide), N-oleoylethanolamine, N-palmitoylethanolamine, 2-arachidonoylglycerol, and its inactive isomer 1-arachidonoylglycerol. On-line sample trapping coupled with separation via microflow liquid chromatography and detection by tandem quadrupole mass spectrometry results in the necessary sensitivity for accurate quantitation of ECs in less than 50μL of serum, without the need for off-line solid phase extraction. Limits of quantitation between 1.2 and 13.4pg/mL were achieved, representing a significant increase in sensitivity compared to previous methods using analytical flow rates. An additional benefit of microflow chromatography is the reduction of solvent consumption by more than two orders of magnitude. The experimental utility of the assay is demonstrated through the analysis of serum from hibernating bears to assess seasonal changes in circulating EC concentrations.

The endocannabinoid anandamide (N-arachidonoylethanolamine, AEA), a physiologically occurring bioactive compound on CB1 and CB2 receptors, has multiple physiological functions. Since the discovery of AEA additional non-cannabinoid endogenous compounds such as N-palmitoylethanolamine (PEA), and N-oleoylethanolamine (OEA) have been identified from mammalian tissues. Virodhamine (O-arachidonoylethanolamine, VA) is the only identified new member of the endocannabinoid family that is characterised by an ester linkage between acylic acid and ethanolamine instead of the amide linkage found in AEA and others non-cannabinoid N-acylethanolamines. It has been reported, as a cautionary note for lipid analyses, that VA can be produced nonenzymatically from AEA (and vice versa) as consequence of O,N-acyl migrations. O,N-acyl migrations are well documented in synthetic organic chemistry literature, but are not well described or recognized with regard to methods in lipid isolation or lipid enzyme studies. We here report an economical and effective protocol for large scale synthesis and characterization of some N- and O-acylethanolamines that could be useful as reference standards in order to investigate their possible formation in biological membranes, with potentially interesting biological properties.

Although N-arachidonoylethanolamine (AEA; also known as anandamide) is present in human follicular fluid (FF), its regulation remains unknown. Therefore, the aims of the present study were to: (1) investigate the relationships between FF AEA concentrations in women undergoing assisted reproductive technology and their age, body mass index, ART characteristics and fertility treatment outcomes; and (2) assess how different inflammatory patterns may trigger AEA production by human granulosa cells (hGCs). FF AEA concentrations were higher in women undergoing IVF than in those undergoing intracytoplasmic sperm injection group. FF AEA median concentrations were lower in women undergoing ART because of male factor infertility than in women with endometriosis (1.6 vs 2.5nM respectively), but not women with tubal, hormonal or unexplained infertility (1.6, 2.4 and 1.9nM respectively). To evaluate the effects of macrophages on AEA production by hGCs, hGCs were cocultured with monocyte-derived macrophages. The conditioned medium from M1 polarised macrophages increased AEA production by hGCs. This was accompanied by an increase in AEA-metabolising enzymes, particularly N-acyl phosphatidylethanolamine-specific phospholipase D. The results of the present study show that high FF AEA concentrations in patients with endometriosis may be associated with the recruitment of inflammatory chemokines within the ovary, which together may contribute to the decreased reproductive potential of women with endometriosis. Collectively, these findings add a new player to the hormone and cytokine networks that regulate fertility in women.

Objective: Endocannabinoids have been implicated in the pathophysiology of Major Depressive Disorder (MDD) and might represent potential targets for therapeutic intervention. Objectives of the study were: (1) to measure plasma levels of endocannabinoids in a group of antidepressant-free depressed outpatients; (2) to explore their relationship with the severity of depressive symptoms as subjectively perceived by the patients; and (3) to investigate the effect of the selective serotonin reuptake inhibitor escitalopram on endocannabinoid levels.

Methods: We measured plasma levels of the two major endocannabinoids, 2-arachidonoylglycerol (2-AG) and N-arachidonoylethanolamine (anadamide), in 12 drug-free outpatients diagnosed with MDD and in 12 matched healthy controls. In the patient group, endocannabinoids plasma levels were assessed at baseline and after 2 months of treatment with escitalopram.

Results: Baseline plasma levels of the two endocannabinoids did not differ between depressed patients and healthy controls. However, there was a significant inverse correlation between 2-arachidonoylglycerol levels and the severity of subjectively perceived depressive symptoms. Treatment with escitalopram did not change endocannabinoid levels in depressed patients, although it caused the expected improvement of depressive symptoms.

Conclusions: Our results suggest that 2-arachidonylglycerol, the most abundant endocannabinoid in the central nervous system, might act to mitigate depressive symptoms, and raise the interesting possibility that 2-arachidonylglycerol and anandamide are differentially regulated in patients affected by MDD. Also, our data suggest but do not prove that the endocannabinoid system is not regulated by serotonergic transmission, at least in depressed patients.

Keywords: 2-AG; AEA; depression; endocannabinoids; escitalopram.

Accumulated evidence suggests that enhancing the endocannabinoid (eCB) tone, in particular of anandamide (N-arachidonoylethanolamine, AEA), has therapeutic potential in many human diseases. Fatty acid amide hydrolase (FAAH) is a membrane-bound enzyme principally responsible for the degradation of AEA, and thus it represents a relevant target to increase signaling thereof. In recent years, different synthetic and natural compounds have been developed and tested on rat FAAH, but little is known of their effect on the human enzyme. Here, we sought to investigate six major cannabis-derived compounds to compare their action on rat and human FAAHs. To this aim, we combined an in silico analysis of their binding mode and affinity, with in vitro assays of their effect on enzyme activity. This integrated approach allowed to disclose differences in efficacy towards rat and human FAAHs, and to highlight the role of key residues involved in the inhibition of both enzymes. This study suggests that the therapeutic efficacy of compounds targeted towards FAAH should be always tested in vitro on both rat and human enzymes.

Keywords: FAAH; cannabinoids; docking; endocannabinoids; inhibition; modeling.

According to the World Health Organization (WHO), 47 million of people display mental health disorders Worldwide. In addition, epidemiological studies have shown that the extension of life expectancy and the increase in aged population will significantly impact in the prevalence of several mental impairments. Although there are strategies for preventing and alleviate mental illnesses such as pharmacological and psychological approaches, limited results have been observed. Thus, the search for new therapeutics for managing psychiatric disorders has explored multiple roads. In recent years, it has been demonstrated that physical activity and exercise promote health benefits. On the other hand, among the neurobiological systems that participate in the genesis and development of mental disruptions, the endocannabinoid system has been suggested as an active player. Supporting this hypothesis, data suggest that the elements comprising the endocannabinoid system, such as the CB1/CB2 cannabinoid receptors, endogenous ligands (N-arachidonoylethanolamine [anandamide, AEA] and 2-arachidonoylglycerol [2- AG]), transporters and the enzymes involved in the biosynthesis and degradation of the AEA and 2-AG, modulate mental diseases. In this review, we discuss that the endocannabinoid system might be considered as a modulator for the positive outcomes of exercise in the management of mental disorders. Clinically, this promising field might be exploited by targeting the elements of the endocannabinoid system aimed to increase the exercise benefits applied to patients with mental illnesses.

Keywords: Anandamide; endocannabinoids; physical activity; mental health..

N-Docosahexaenoylethanolamine (synaptamide) is structurally similar to the endocannabinoid N-arachidonoylethanolamine (anandamide), but incorporates the omega-3 22:6 fatty acid docosahexaenoic acid (DHA) in place of the omega-6 20:4 fatty acid arachidonic acid (AA). Some brain membrane lipid effects may be mediated via synaptamide. In competition experiments with mouse brain homogenate in vitro, we found that synaptamide was an order-of-magnitude poorer inhibitor of radioactive anandamide hydrolysis than was anandamide itself. Also, enzyme-mediated hydrolysis of synaptamide was observed to occur at a slower rate than for anandamide. We have synthesized synaptamide radiolabeled with carbon-14 in both the ethanolamine ([α,β-14C2]synaptamide) and in the DHA ([1-14C]synaptamide) moieties. The brain penetration, distribution, and metabolism of radiolabeled synaptamide were studied in mice in vivo relative to anandamide, DHA, and AA. Brain uptake of labeled synaptamide was greater than for labeled DHA, consistent with previous studies of labeled anandamide and AA in our laboratory. After administering either isotopomer of radiolabeled synaptamide, radiolabeled phospholipids were found in mouse brain. Pretreatment of mice with PF3845, a potent, specific inhibitor of fatty acid amide hydrolase (FAAH), eliminated formation of labeled phospholipids measured after 15min, suggesting that synaptamide is hydrolyzed nearly exclusively by FAAH, though it is a poorer substrate for FAAH than anandamide.

Human immunodeficiency virus (HIV) infection and antiretroviral therapy can independently induce HIV-associated neuropathic pain (HIV-NP). There is a dearth of drugs or therapeutic modalities that can alleviate HIV-NP. Smoked cannabis has been reported to improve pain measures in patients with neuropathic pain. Cannabis, phytocannabinoids, and the endocannabinoids such N-arachidonoylethanolamine (anandamide; AEA) and 2-arachidonoylglycerol (2-AG), produce some of their effects via cannabinoid receptors (CBRs). Endocannabinoids are degraded by various enzymes such as fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase. We searched PubMed, Google Scholar, clinicaltrials.gov and clinicaltrialsregister.eu using various key words and their combinations for published papers that studied HIV-NP and cannabis, cannabinoids, or endocannabinoids up to 27th December 2020. All original research articles that evaluated the efficacy of molecules that modulate the endocannabinoid system (ECS) for the prevention and/or treatment of pain in HIV-NP animal models and patients with HIV-NP were included. The PubMed search produced a total of 117 articles, whereas the Google Scholar search produced a total of 9467 articles. Amongst the 13 articles that fulfilled the inclusion criteria 11 articles were found in both searches whereas 2 articles were found in Google Scholar only. The clinicaltrials.gov and clinicaltrialsregister.eu searches produced five registered trials of which three were completed and with results. Ten preclinical studies found that the endocannabinoids (2-AG and AEA), synthetic mixed CB1R/CB2R agonist WIN 55,212-2, a CB2R-selective phytocannabinoid β-caryophyllene, synthetic CB2R-selective agonists (AM1710, JWH015, JWH133 and Gp1a, but not HU308); FAAH inhibitors (palmitoylallylamide, URB597 and PF-3845) and a drug combination of indomethacin plus minocycline, which produces its effects in a CBR-dependent manner, either prevented the development of and/or attenuated established HIV-NP. Two clinical trials demonstrated greater efficacy of smoked cannabis over placebo in alleviating HIV-NP, whereas another clinical trial demonstrated that cannabidivarin, a cannabinoid that does not activate CBRs, did not reduce HIV-NP. The available preclinical results suggest that targeting the ECS for prevention and treatment of HIV-NP is a plausible therapeutic option. Clinical evidence shows that smoked cannabis alleviates HIV-NP. Further research is needed to find out if non-psychoactive drugs that target the ECS and are delivered by other routes than smoking could be useful as treatment options for HIV-NP.

Keywords: 2-AG, 2-arachidonoylglycerol; ABHD12, α-β-hydrolase domain-containing 12; ABHD6, α-β-hydrolase domain-containing 6; AEA, N-arachidonoylethanolamine; AEs, adverse effects; Antiretroviral; BCP, β-caryophyllene; CB1R, cannabinoid type 1 receptor; CB2R, cannabinoid type 2 receptor; CBD, cannabidiol; CBDV, cannabidivarin; CBRs, cannabinoid receptors; CINP, chemotherapy-induced neuropathic pain; CNS, central nervous system; COX, cyclooxygenase; Cannabinoid; Cannabis; DAG, diacylglycerol; DAGL, DAG lipase; DDS, descriptor differential scale; DSP, distal symmetric polyneuropathy; ECS, endocannabinoid system; Endocannabinoid; FAAH, fatty acid amide hydrolase; FDA, Food and Drug Administration; GPCRs, G protein-coupled receptors; HIV, human immunodeficiency virus; HIV-DSP, HIV-distal symmetric polyneuropathy; HIV-NP, HIV-associated neuropathic pain; Human immunodeficiency virus; IPM, indomethacin plus minocycline; L-29, palmitoylallylamide; MAGL, monoacylglycerol lipase; MAIDS, murine acquired immunodeficiency syndrome; NAPE, N-acyl-phosphatidylethanolamine; NAPE-PLD, NAPE-specific phospholipase D; NP, neuropathic pain; NSAIDs, non-steroidal anti-inflammatory drugs; Neuropathic pain; OTC, over the counter; PLWH, people living with HIV; PNP, peripheral neuropathic pain; RCTs, randomised clinical trials; SAMRC, South African Medical Research Council; TRPA, transient receptor potential ankyrin; TRPV, transient receptor potential vanilloid; WHO, World Health Organization; ddC, 2′-3′-dideoxycytidine; delta-9-THC, delta-9-tetrahydrocannabinol; gp, glycoprotein.

Δ9-tetrahydrocannabinol (Δ9-THC), the main active ingredient of Cannabis sativa (marijuana), interacts with the human brain cannabinoid (CB1) receptor and mimics pharmacological effects of endocannabinoids (eCBs) like N-arachidonylethanolamide (AEA). Due to its flexible nature of AEA structure with more than 15 rotatable bonds, establishing its binding mode to the CB1 receptor is elusive. The aim of the present study was to explore possible binding conformations of AEA within the binding pocket of the CB1 receptor confirmed in the recently available X-ray crystal structures of the CB1 receptor and predict essential AEA binding domains. We performed long time molecular dynamics (MD) simulations of plausible AEA docking poses until its receptor binding interactions became optimally established. Our simulation results revealed that AEA favors to bind to the hydrophobic channel (HC) of the CB1 receptor, suggesting that HC holds essential significance in AEA binding to the CB1 receptor. Our results also suggest that the Helix 2 (H2)/H3 region of the CB1 receptor is an AEA binding subsite privileged over the H7 region.

Endocannabinoids are a group of endogenous mediators derived from membrane lipids, which are implicated in a wide variety of physiological functions such as blood pressure regulation, immunity, pain, memory, reward, perception, reproduction, and sleep. N-Arachidonoylethanolamine (anandamide; AEA) and 2-arachidonoylglycerol (2-AG) represent two major endocannabinoids in the human body and they exert many of their cellular and organ system effects by activating the G_i/o protein-coupled, cannabinoid type 1 (CB1) and type 2 (CB2) receptors. However, not all effects of cannabinoids are ascribable to their interaction with CB1 and CB2 receptors; indeed, macromolecules like other types of receptors, ion channels, transcription factors, enzymes, transporters, and cellular structure have been suggested to mediate the functional effects of cannabinoids. Among the proposed molecular targets of endocannabinoids, potassium channels constitute an intriguing group, because these channels not only are crucial in shaping action potentials and controlling the membrane potential and cell excitability, thereby regulating a wide array of physiological processes, but also serve as potential therapeutic targets for the treatment of cancer and metabolic, neurological and cardiovascular disorders. This review sought to survey evidence pertaining to the CB1 and CB2 receptor-independent actions of endocannabinoids on ion channels, with an emphasis on AEA and potassium channels. To better understand the functional roles as well as potential medicinal uses of cannabinoids in human health and disease, further mechanistic studies to delineate interactions between various types of cannabinoids and ion channels, including members in the potassium channel superfamily, are warranted.

Keywords: 2-arachidonoylglycerol (2-AG); Potassium channels; anandamide (AEA); endocannabinoid system; non-CB1, non-CB2 receptor-mediated.

Introduction: The endocannabinoid system (ECS) regulates functions throughout human physiology, including neuropsychiatric, cardiovascular, autonomic, metabolic, and inflammatory states. The complex cellular interactions regulated by the ECS suggest a potential for vascular disease and stroke prevention by augmenting central nervous and immune cell endocannabinoid signaling. Discussion: The endocannabinoid N-arachidonoylethanolamine (anandamide) plays a central role in augmenting these processes in cerebrovascular and neurometabolic disease. Furthermore, cannabidiol (CBD), a nonpsychoactive constituent of Cannabis, is an immediate therapeutic candidate both for potentiating endocannabinoid signaling and for acting at multiple pharmacological targets. Conclusion: This speculative synthesis explores the current state of knowledge of the ECS and suggests CBD as a therapeutic candidate for stroke prevention by exerting favorable augmentation of the homeostatic effects of the ECS and, in turn, improving the metabolic syndrome, while simultaneously stalling the development of atherosclerosis.

The endocannabinoids are now known as novel and important regulators of energy metabolism and homeostasis. The endocrine functions of white adipose are chiefly involved in the control of whole-body metabolism, insulin sensitivity and food intake. Adipocytes produce hormones, such as leptin and adiponectin, that can improve insulin resistance or peptides, such as TNF-α, that elicit insulin resistance. Adipocytes express specific receptors, such as peroxisome proliferator-activated receptor (PPAR)-γ, which serve as adipocyte targets for insulin sensitizers such as thiazolidinediones. Recently, endocannabinoids and related compounds were identified in human fat cells. The endocannabinoid system consists primarily of two receptors, cannabinoid (CB)1 and CB2, their endogenous ligands termed endocannabinoids and the enzymes responsible for ligand biosynthesis and degradation. The endocannabinoids 2-arachidonylglycerol and anandamide or N-arachidonoylethanolamine increase food intake and promote weight gain in animals. Rimonabant, a selective CB1 blocker, reduces food intake and body weight in animals and humans.

The endocannabinoid system is involved in the regulation of the stress response, but the relative contribution of N-arachidonoylethanolamine (AEA) and 2-arachidonoylglycerol (2-AG) and their mechanisms have to be elucidated. In this study, we compared the effects of the pharmacological inhibition of the two major endocannabinoid-degrading enzymes [fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL) for AEA and 2-AG, respectively] on stress-coping [forced swim test (FST) and tail suspension test (TST)] and anxiety-like [elevated-plus maze (EPM) and light-dark test (LDT)] behaviors in wild-type and FAAH knockout mice. In vivo microdialysis estimated the effects of FAAH and MAGL inhibition on dopamine (DA) and serotonin (5-HT) levels in the medial prefrontal cortex (mPFC) during an FST. Mice were treated with PF-3845 (FAAH inhibitor), JZL184 (MAGL inhibitor), JZL195 (dual FAAH/MAGL inhibitor) or vehicle. Our data showed that PF-3845 increased latency to immobility and decreased total immobility time in FST, but no effects were observed in TST compared with vehicle-treated wild-type mice. By contrast, JZL184 decreased latency and increased immobility in TST and FST. JZL195 in wild-type mice and JZL184 in FAAH knockout mice reproduced the same passive coping behaviors as JZL184 in wild-type mice in TST and FST. In the microdialysis experiment, FST was associated with increased DA and 5-HT levels in the mPFC. However, JZL184-treated wild-type mice displayed a significant attenuation of forced swim stress-induced DA release compared with vehicle-treated wild-type mice and PF-3845-treated wild-type mice. Finally, FAAH and/or MAGL inhibitors induced robust and consistent anxiolytic-like effects in EPM and LDT. These results suggested differences between FAAH and MAGL inhibition in stress-coping behaviors. Notably, MAGL inhibition induced a consistent avoidant coping behavior and attenuated the stress-induced mPFC DA response in FST. However, more investigation is needed to elucidate the functional association between DA and 2-AG signaling pathways, and the molecular mechanism in the regulation of passive coping strategies during inescapable stress.

Keywords: 2-Arachidonoylglycerol; Dopamine; Microdialysis; Mouse; Stress-coping behavior.

Alzheimer's disease (AD) is characterized by multiple cognitive deficits including memory and sensorimotor gating impairments as a result of neuronal and synaptic loss. The endocannabinoid system plays an important role in these deficits but little is known about its influence on the molecular mechanism regarding phosphorylated tau (p-tau) protein accumulation - one of the hallmarks of AD -, and on the density of synaptic proteins. Thus, the aim of this study was to investigate the preventive effects of anandamide (N-arachidonoylethanolamine, AEA) on multiple cognitive deficits and on the levels of synaptic proteins (syntaxin 1, synaptophysin and synaptosomal-associated protein, SNAP-25), cannabinoid receptor type 1 (CB1) and molecules related to p-tau degradation machinery (heat shock protein 70, HSP70), and Bcl2-associated athanogene (BAG2) in an AD-like sporadic dementia model in rats using intracerebroventricular (icv) injection of streptozotocin (STZ). Our hypothesis is that AEA could interact with HSP70, modulating the level of p-tau and synaptic proteins, preventing STZ-induced cognitive impairments. Thirty days after receiving bilateral icv injections of AEA or STZ or both, the cognitive performance of adult male Wistar rats was evaluated in the object recognition test, by the escape latency in the elevated plus maze (EPM), by the tone and context fear conditioning as well as in prepulse inhibition tests. Subsequently, the animals were euthanized and their brains were removed for histological analysis or for protein quantification by Western Blotting. The behavioral results showed that STZ impaired recognition, plus maze and tone fear memories but did not affect contextual fear memory and prepulse inhibition. Moreover, AEA prevented recognition and non-associative emotional memory impairments induced by STZ, but did not influence tone fear conditioning. STZ increased the brain ventricular area and this enlargement was prevented by AEA. Additionally, STZ reduced the levels of p-tau (Ser199/202) and increased p-tau (Ser396), although AEA did not affect these alterations. HSP70 was found diminished only by STZ, while BAG2 levels were decreased by STZ and AEA. Synaptophysin, syntaxin and CB1 receptor levels were reduced by STZ, but only syntaxin was recovered by AEA. Altogether, albeit AEA failed to modify some AD-like neurochemical alterations, it partially prevented STZ-induced cognitive impairments, changes in synaptic markers and ventricle enlargement. This study showed, for the first time, that the administration of an endocannabinoid can prevent AD-like effects induced by STZ, boosting further investigations about the modulation of endocannabinoid levels as a therapeutic approach for AD.

<stro<em>n</em>g class="sub-title"> I<em>n</em>troductio<em>n</em>: </stro<em>n</em>g> Several drugs of abuse (DOA) are capable of modulati<em>n</em>g <em>n</em>eurohypophysial hormo<em>n</em>es, such as oxytoci<em>n</em> (OT) a<em>n</em>d vasopressi<em>n</em> (VP), pote<em>n</em>tially resulti<em>n</em>g i<em>n</em> the developme<em>n</em>t of psychological ab<em>n</em>ormalities, such as cog<em>n</em>itive dysfu<em>n</em>ctio<em>n</em>, psychoses, a<em>n</em>d affective disorders. Efavire<em>n</em>z (EFV), widely used i<em>n</em> Africa a<em>n</em>d globally to treat HIV, i<em>n</em>duces diverse <em>n</em>europsychiatric side effects while its abuse has become a global co<em>n</em>cer<em>n</em>. The actio<em>n</em>s of EFV may i<em>n</em>volve <em>n</em>eurohypophysial system (<em>N</em>S) disruptio<em>n</em> like that of k<em>n</em>ow<em>n</em> DOA. This study i<em>n</em>vestigated whether sub-chro<em>n</em>ic EFV exposure, at a previously-determi<em>n</em>ed rewardi<em>n</em>g dose, alters peripheral OT a<em>n</em>d VP levels versus that of a co<em>n</em>trol, ∆⁹-tetrahydroca<em>n</em><em>n</em>abi<em>n</em>ol (∆⁹-THC), methamphetami<em>n</em>e (MA) a<em>n</em>d cocai<em>n</em>e.

<stro<em>n</em>g class="sub-title"> Materials a<em>n</em>d methods: </stro<em>n</em>g> To simulate the co<em>n</em>ditio<em>n</em>s u<em>n</em>der which reward-drive<em>n</em> behavior had previously bee<em>n</em> established for EFV, male Sprague Dawley rats (<i><em>n</em></i> = 16/exposure) received i<em>n</em>traperito<em>n</em>eal vehicle (co<em>n</em>trol) or drug admi<em>n</em>istratio<em>n</em> across a<em>n</em> alter<em>n</em>ati<em>n</em>g sixtee<em>n</em>-day dosi<em>n</em>g protocol. Co<em>n</em>trol admi<em>n</em>istratio<em>n</em> (sali<em>n</em>e/olive oil; 0.2 ml) occurred o<em>n</em> odd-<em>n</em>umbered a<em>n</em>d drug admi<em>n</em>istratio<em>n</em> (EFV: 5 mg/kg, ∆⁹-THC: 0.75 mg/kg, MA: 1 mg/kg, or cocai<em>n</em>e: 20 mg/kg) o<em>n</em> eve<em>n</em>-<em>n</em>umbered days followed by eutha<em>n</em>asia, tru<em>n</em>k blood collectio<em>n</em> a<em>n</em>d plasma extractio<em>n</em> for <em>n</em>europeptide assay. Effect of drug exposure o<em>n</em> peripheral OT a<em>n</em>d VP levels was assessed versus co<em>n</em>trols a<em>n</em>d qua<em>n</em>tified usi<em>n</em>g specific ELISA kits. Statistical sig<em>n</em>ifica<em>n</em>ce was determi<em>n</em>ed by Kruskal-Wallis A<em>N</em>OVA, with p < 0.05. Ethics approval: <em>N</em>WU-00291-17-A5.

<stro<em>n</em>g class="sub-title"> Results: </stro<em>n</em>g> Delta-9-THC reduced OT a<em>n</em>d VP plasma levels (<i>p</i> < 0.0001, <i>p</i> = 0.0141; respectively), cocai<em>n</em>e reduced plasma OT (<i>p</i> = 0.0023), while MA reduced plasma VP levels (<i>p</i> = 0.0001), all versus co<em>n</em>trol. EFV reduced OT a<em>n</em>d VP plasma levels (<i>p</i> < 0.0001; OT a<em>n</em>d VP) versus co<em>n</em>trol, a<em>n</em>d similar to ∆⁹-THC.

<stro<em>n</em>g class="sub-title"> Co<em>n</em>clusio<em>n</em>: </stro<em>n</em>g> EFV markedly affects the <em>N</em>S i<em>n</em> sig<em>n</em>ifica<em>n</em>tly reduci<em>n</em>g <i>both</i> plasma OT a<em>n</em>d VP equivale<em>n</em>t to DOA. Importa<em>n</em>tly, EFV has disti<em>n</em>ct effects o<em>n</em> peripheral OT a<em>n</em>d VP levels whe<em>n</em> assessed withi<em>n</em> the co<em>n</em>text of drug depe<em>n</em>de<em>n</em>ce. The data highlights a possible <em>n</em>ew mecha<em>n</em>ism u<em>n</em>derlyi<em>n</em>g previously docume<em>n</em>ted EFV-i<em>n</em>duced effects i<em>n</em> rats, a<em>n</em>d whereby EFV may i<em>n</em>duce <em>n</em>europsychiatric adverse effects cli<em>n</em>ically; also providi<em>n</em>g a deeper u<em>n</em>dersta<em>n</em>di<em>n</em>g of the suggested abuse-pote<em>n</em>tial of EFV.

<stro<em>n</em>g class="sub-title"> Keywords: </stro<em>n</em>g> 5-HT, 5-hydroxytryptami<em>n</em>e (seroto<em>n</em>i<em>n</em>); ADH, a<em>n</em>tidiuretic hormo<em>n</em>e; AEA, <em>N</em>-<em>arachidonoylethanolamine</em> (a<em>n</em>a<em>n</em>damide); A<em>N</em>OVA, o<em>n</em>e-way a<em>n</em>alysis of varia<em>n</em>ce; ARRIVE, a<em>n</em>imal research: reporti<em>n</em>g of i<em>n</em> vivo experime<em>n</em>ts (guideli<em>n</em>es); ARV, a<em>n</em>tiretroviral; Ach, acetylcholi<em>n</em>e; CB, ca<em>n</em><em>n</em>abi<em>n</em>oid; C<em>N</em>S, ce<em>n</em>tral <em>n</em>ervous system; CPP, co<em>n</em>ditio<em>n</em>ed place prefere<em>n</em>ce; Cocai<em>n</em>e; DA, dopami<em>n</em>e; DAT, dopami<em>n</em>e tra<em>n</em>sporter; DOA‘s, drug(s) of abuse; ECS, e<em>n</em>doca<em>n</em><em>n</em>abi<em>n</em>oid system; EFV, efavire<em>n</em>z; ELISA, e<em>n</em>zyme-li<em>n</em>ked immu<em>n</em>osorbe<em>n</em>t assay; Efavire<em>n</em>z; GABA, gamma-ami<em>n</em>obutyric acid; Glu, glutamate; HIV, huma<em>n</em> immu<em>n</em>odeficie<em>n</em>cy virus; H<em>N</em>S, hypothalamic <em>n</em>eurohypophysial system; HPA, hypothalamic-pituitary-adre<em>n</em>al (axis); IP, i<em>n</em>traperito<em>n</em>eal; IV, i<em>n</em>trave<em>n</em>ous; M, muscari<em>n</em>ic; MA, methamphetami<em>n</em>e; MAO, mo<em>n</em>oami<em>n</em>e oxidase; Methamphetami<em>n</em>e; <em>N</em>Ac, <em>n</em>ucleus accumbe<em>n</em>s; <em>N</em>E, <em>n</em>orepi<em>n</em>ephri<em>n</em>e; <em>N</em>O, <em>n</em>itric oxide; <em>N</em>PAE, <em>n</em>europsychiatric adverse effect; OT, oxytoci<em>n</em>; OTR, oxytoci<em>n</em> receptor; Oxytoci<em>n</em>; P<em>N</em>D, post<em>n</em>atal day; PV<em>N</em>, parave<em>n</em>tricular <em>n</em>ucleus; SC, subcuta<em>n</em>eous; SD, Sprague Dawley (rat); SEM, sta<em>n</em>dard error of the mea<em>n</em>; SERT, seroto<em>n</em>i<em>n</em> tra<em>n</em>sporter; SO<em>N</em>, supraoptic <em>n</em>ucleus; VMAT, vesicular mo<em>n</em>oami<em>n</em>e tra<em>n</em>sporter; VP, vasopressi<em>n</em>; VPR, vasopressi<em>n</em> receptor; Vasopressi<em>n</em>; cART, combi<em>n</em>ed a<em>n</em>tiretroviral therapy; ∆9-THC, delta-9-tetrahydroca<em>n</em><em>n</em>abi<em>n</em>ol; ∆9-tetrahydroca<em>n</em><em>n</em>abi<em>n</em>ol.

Cannabinoids, including cannabis derived phytocannabinoids and endogenous cannabinoids (endocannabinoids), are typically considered anti-inflammatory. One such endocannabinoid is N-arachidonoylethanolamine (anandamide, AEA), which is metabolized by fatty acid amide hydrolase (FAAH). In humans, there is a loss of function single nucleotide polymorphism (SNP) in the FAAH gene (C385A, rs324420), that leads to increases in the levels of AEA. Using a mouse model with this SNP, we investigated how this SNP affects inflammation in a model of inflammatory bowel disease. We administered 2,4,6-trinitrobenzene sulfonic acid (TNBS) intracolonically, to adult male FAAH SNP mice and examined colonic macroscopic tissue damage and myeloperoxidase activity, as well as levels of plasma and amygdalar cytokines and chemokines 3 days after administration, at the peak of colitis. We found that mice possessing the loss of function alleles (AC and AA), displayed no differences in colonic damage or myeloperoxidase activity compared to mice with wild type alleles (CC). In contrast, in plasma, colitis-induced increases in interleukin (IL)-2, leukemia inhibitory factor (LIF), monocyte chemoattractant protein (MCP)-1, and tumor necrosis factor (TNF) were reduced in animals with an A allele. A similar pattern was observed in the amygdala for granulocyte colony stimulating factor (G-CSF) and MCP-1. In the amygdala, the mutant A allele led to lower levels of IL-1α, IL-9, macrophage inflammatory protein (MIP)-1β, and MIP-2 independent of colitis-providing additional understanding of how FAAH may serve as a regulator of inflammatory responses in the brain. Together, these data provide insights into how FAAH regulates inflammatory processes in disease.

Keywords: amygdala; colitis; cytokines; endocannabinoids; inflammation.

The use of cannabis for skin diseases and hair regrowth is at the preliminary stage.

Legalization: Many countries have approved cannabis for medical use; however, four countries Canada, Uruguay, South Africa, and Georgia have legalized it for both medical and recreational purposes.

The endocannabinoid system: The endocannabinoid system may maintain skin homeostasis; two notable endocannabinoids include 2-Arachidonoylglycerol (2-AG) and N-arachidonoylethanolamine (AEA).

Routes of administration and pharmacokinetics: Topical cannabinoids can avoid the first-pass metabolism and reduce respiratory side effects; however, the high hydrophobicity of cannabinoids may hinder percutaneous absorption.

Skin disorders and hair growth: Human clinical studies suggest that cannabinoids may be used in eczema, acne, pruritus, and systemic sclerosis treatment. Cannabidiol (CBD) may enhance hair growth via multiple mechanisms.

Safety: Topical cannabis may cause mild side effects such as pruritus, burning, erythema, and stinging; they are relatively safer than inhalation and oral cannabis. Cannabis use may be associated with allergic symptoms and reduced immune response to live vaccination.

Cannabinoids in practice: Despite growing interest, dermatologists should be cautious prescribing cannabinoids due to insufficient clinical data on both efficacy and safety.

Keywords: cannabidiol; cannabinoids; endocannabinoid system; hair growth; skin diseases.

The ability of endocannabinoid (eCB) to change functional microglial phenotype can be explored as a possible target for therapeutic intervention. Since the inhibition of fatty acid amide hydrolase (FAAH), the main catabolic enzyme of anandamide (AEA), may provide beneficial effects in mice model of Alzheimer's disease (AD)-like pathology, we aimed at determining whether the FAAH inhibitor URB597 might target microglia polarization and alter the cytoskeleton reorganization induced by the amyloid-β peptide (Aβ). The morphological evaluation showed that Aβ treatment increased the surface area of BV-2 cells, which acquired a flat and polygonal morphology. URB597 treatment partially rescued the control phenotype of BV-2 cells when co-incubated with Aβ. Moreover, URB597 reduced both the increase of Rho protein activation in Aβ-treated BV-2 cells and the Aβ-induced migration of BV-2 cells, while an increase of Cdc42 protein activation was observed in all samples. URB597 also increased the number of BV-2 cells involved in phagocytosis. URB597 treatment induced the polarization of microglial cells towards an anti-inflammatory phenotype, as demonstrated by the decreased expression of iNOS and pro-inflammatory cytokines along with the parallel increase of Arg-1 and anti-inflammatory cytokines. Taken together, these data suggest that FAAH inhibition promotes cytoskeleton reorganization, regulates phagocytosis and cell migration processes, thus driving microglial polarization towards an anti-inflammatory phenotype.

Keywords: M1/M2 phenotypes; N-arachidonoylethanolamine (AEA); URB597; actin cytoskeleton; cell migration; fatty acid amide hydrolase (FAAH); microglia; neuroinflammation.

The endocannabinoid system is involved in the regulation of a variety of physiological and cognitive processes. While the endocannabinoids 2-arachidonoylglycerol (2-AG) and anandamide (N-arachidonoylethanolamine, AEA) have been found in breast milk, their role(s) have yet to be determined. This study determined the normal concentration ranges of endocannabinoids (2-AG and AEA) in breast milk and the influences, if any, of obesity and diurnal rhythms on their levels. Milk samples were collected from 36 breastfeeding mothers at 4-8 weeks postpartum at each feed over a 24-h period, and further stratified into three groups based on body mass index (BMI). The samples were analyzed using liquid chromatography mass spectrometry. AEA was below the limit of detection and 2-AG levels averaged 59.3 ± 18.3 ng/mL (± SD) in women with normal BMI. Wide-ranging 2-AG concentrations in the overweight (65.5 ± 41.9 ng/mL) /obese (66.1 ± 40.6 ng/mL) groups suggest BMI may be a contributing factor influencing its levels. Following a diurnal pattern, there was a significantly higher 2-AG concentration observed during the day, as compared to night time samples. In conclusion, our study clearly suggests that appropriate milk collection and storage conditions are critical. Further, body weight and diurnal rhythm appear to influence levels of 2-AG. Based on these results, future studies are underway to determine what specific roles endocannabinoids may play in human milk and how elevated levels of 2-AG may modulate infant appetite and health.

Keywords: 2-arachidonoylglycerol; anandamide; diurnal; endocannabinoids; human milk; obesity.

Background: Loneliness is one of the most distressing grief symptoms and is associated with adverse mental health in bereaved older adults. The endocannabinoid signaling (ECS) system is stress-responsive and circulating endocannabinoid (eCB) concentrations are elevated following bereavement. This study examined the association between loneliness and circulating eCB concentrations in grieving older adults and explored the role of eCBs on the association between baseline loneliness and grief symptom trajectories. Methods: A total of 64 adults [grief with high loneliness: n = 18; grief with low loneliness: n = 26; and healthy comparison (HC): n = 20] completed baseline clinical assessments for the UCLA loneliness scale. In grief participants, longitudinal clinical assessments, including the Inventory of Complicated Grief and 17-item Hamilton Depression Rating scales, were collected over 6 months. Baseline circulating eCB [N-arachidonoylethanolamine (AEA) and 2-arachidonoylglycerol (2-AG)] concentrations were quantified in the serum using isotope dilution, liquid chromatography-mass spectrometry; cortisol concentrations were measured in the same samples using radioimmunoassay. Results: Circulating AEA concentrations were higher in severely lonely grieving elders than in HC group; cortisol concentrations were not different among the groups. Cross-sectionally, loneliness scores were positively associated with AEA concentrations in grievers; this finding was not significant after accounting for depressive symptom severity. Grieving individuals who endorsed high loneliness and had higher 2-AG concentrations at baseline showed faster grief symptom resolution. Conclusions: These novel findings suggest that in lonely, bereaved elders, increased circulating eCBs, a reflection of an efficient ECS system, are associated with better adaptation to bereavement. Circulating eCBs as potential moderators and mediators of the loneliness-grief trajectory associations should be investigated.

<stro<em>n</em>g class="sub-title"> Keywords: </stro<em>n</em>g> 2-arachido<em>n</em>oylglycerol; <em>N</em>-<em>arachidonoylethanolamine</em>; bereaveme<em>n</em>t; e<em>n</em>doca<em>n</em><em>n</em>abi<em>n</em>oid; grief; lo<em>n</em>eli<em>n</em>ess; older adult; prolo<em>n</em>ged grief disorder (PGD).

This article summarizes our early work with Viswanathan Natarajan in the 1980s at the University of Minnesota's Hormel Institute, when he was at the beginning of his brilliant academic career. At that time most metabolic pathways for the biosynthesis and degradation of phospholipids were well established and known in considerable detail. Hence, it was exciting to discover a novel sequence of biochemical reactions, first in dog heart and later in various other vertebrate cells and tissues that became known as the transacylation-phosphodiesterase pathway of phospholipid metabolism. Because one of the metabolites, N-arachidonoylethanolamine, produced by this reaction sequence, was later found to bind to and activate cannabinoid receptors, investigations of this pathway became part of the rapidly growing field of endocannabinoid research. This is briefly summarized here as well.