A Delightful Trip Along the Pathway of Cannabinoid and Endocannabinoid Chemistry and Pharmacology.

After a traumatic childhood in Europe during the Second World War, I found that scientific research in Israel was a pleasure beyond my expectations. Over the last 65 year, I have worked on the chemistry and pharmacology of natural products. During the last few decades, most of my research has been on plant cannabinoids, the endogenous cannabinoids arachidonoyl ethanolamide (anandamide) and 2-arachidonoyl glycerol, and endogenous anandamide-like compounds, all of which are involved in a wide spectrum of physiological reactions. Two plant cannabinoids, Δ9-tetrahydrocannabinol and cannabidiol, are approved drugs. However, the endogenous cannabinoids and the anandamide-like constituents have not yet been well investigated in humans. For me, intellectual freedom-the ability to do research based on my own scientific interests-has been the most satisfying part of my working life. Looking back over the 91 years of my long life, I conclude that I have been lucky, very lucky, both personally and scientifically.

Synthetic Cannabinoids: A Pharmacological and Toxicological Overview.

Synthetic cannabinoids (SCs) are a chemically diverse group of new psychoactive substances (NPSs) that target the endocannabinoid system, triggering a plethora of actions (e.g., elevated mood sensation, relaxation, appetite stimulation) that resemble, but are more intense than, those induced by cannabis. Although some of these effects have been explored for therapeutic applications, anticipated stronger psychoactive effects than cannabis and reduced risk perception have increased the recreational use of SCs, which have dominated the NPS market in the United States and Europe over the past decade. However, rising SC-related intoxications and deaths represent a major public health concern and embody a major challenge for policy makers. Here, we review the pharmacology and toxicology of SCs. A thorough characterization of SCs' pharmacodynamics and toxicodynamics is important to better understand the main mechanisms underlying acute and chronic effects of SCs, interpret the clinical/pathological findings related to SC use, and improve SC risk awareness.

SGIP1 binding to the α-helical H9 domain of cannabinoid receptor 1 promotes axonal surface expression.

Endocannabinoid signalling mediated by cannabinoid receptor 1 (CB1R, also known as CNR1) is critical for homeostatic neuromodulation of both excitatory and inhibitory synapses. This requires highly polarised axonal surface expression of CB1R, but how this is achieved remains unclear. We previously reported that the α-helical H9 domain in the intracellular C terminus of CB1R contributes to axonal surface expression by an unknown mechanism. Here, we show in rat primary neuronal cultures that the H9 domain binds to the endocytic adaptor protein SGIP1 to promote CB1R expression in the axonal membrane. Overexpression of SGIP1 increases CB1R axonal surface localisation but has no effect on CB1R lacking the H9 domain (CB1RΔH9). Conversely, SGIP1 knockdown reduces axonal surface expression of CB1R but does not affect CB1RΔH9. Furthermore, SGIP1 knockdown diminishes CB1R-mediated inhibition of presynaptic Ca2+ influx in response to neuronal activity. Taken together, these data advance mechanistic understanding of endocannabinoid signalling by demonstrating that SGIP1 interaction with the H9 domain underpins axonal CB1R surface expression to regulate presynaptic responsiveness.

Cholinergic Neurotransmission Controls Orexigenic Endocannabinoid Signaling in the Gut in Diet-Induced Obesity.

The brain bidirectionally communicates with the gut to control food intake and energy balance, which becomes dysregulated in obesity. For example, endocannabinoid (eCB) signaling in the small-intestinal (SI) epithelium is upregulated in diet-induced obese (DIO) mice and promotes overeating by a mechanism that includes inhibiting gut-brain satiation signaling. Upstream neural and molecular mechanism(s) involved in overproduction of orexigenic gut eCBs in DIO, however, are unknown. We tested the hypothesis that overactive parasympathetic signaling at the muscarinic acetylcholine receptors (mAChRs) in the SI increases biosynthesis of the eCB, 2-arachidonoyl-sn-glycerol (2-AG), which drives hyperphagia via local CB1Rs in DIO. Male mice were maintained on a high-fat/high-sucrose Western-style diet for 60 d, then administered several mAChR antagonists 30 min prior to tissue harvest or a food intake test. Levels of 2-AG and the activity of its metabolic enzymes in the SI were quantitated. DIO mice, when compared to those fed a low-fat/no-sucrose diet, displayed increased expression of cFos protein in the dorsal motor nucleus of the vagus, which suggests an increased activity of efferent cholinergic neurotransmission. These mice exhibited elevated levels of 2-AG biosynthesis in the SI, that was reduced to control levels by mAChR antagonists. Moreover, the peripherally restricted mAChR antagonist, methylhomatropine bromide, and the peripherally restricted CB1R antagonist, AM6545, reduced food intake in DIO mice for up to 24 h but had no effect in mice conditionally deficient in SI CB1Rs. These results suggest that hyperactivity at mAChRs in the periphery increases formation of 2-AG in the SI and activates local CB1Rs, which drives hyperphagia in DIO.

Pre- versus Post-synaptic Forms of LTP in Two Branches of the Same Hippocampal Afferent.

There has been considerable controversy about pre- versus postsynaptic expression of memory-related long-term potentiation (LTP), with corresponding disputes about underlying mechanisms. We report here an instance in male mice, in which both types of potentiation are expressed but in separate branches of the same hippocampal afferent. Induction of LTP in the dentate gyrus (DG) branch of the lateral perforant path (LPP) reduces paired-pulse facilitation, is blocked by antagonism of cannabinoid receptor type 1, and is not affected by suppression of postsynaptic actin polymerization. These observations are consistent with presynaptic expression. The opposite pattern of results was obtained in the LPP branch that innervates the distal dendrites of CA3: LTP did not reduce paired-pulse facilitation, was unaffected by the cannabinoid receptor blocker, and required postsynaptic actin filament assembly. Differences in the two LPP termination sites were also noted for frequency facilitation of synaptic responses, an effect that was reproduced in a two-step simulation by small adjustments to vesicle release dynamics. These results indicate that different types of glutamatergic neurons impose different forms of filtering and synaptic plasticity on their afferents. They also suggest that inputs are routed to, and encoded by, different sites within the hippocampus depending upon the pattern of activity arriving over the parent axon.

Endocannabinoid-Based Therapies.

The endocannabinoids are lipid-derived messengers that play a diversity of regulatory roles in mammalian physiology. Dysfunctions in their activity have been implicated in various disease conditions, attracting attention to the endocannabinoid system as a possible source of therapeutic drugs. This signaling complex has three components: the endogenous ligands, anandamide and 2-arachidonoyl-sn-glycerol (2-AG); a set of enzymes and transporters that generate, eliminate, or modify such ligands; and selective cell surface receptors that mediate their biological actions. We provide an overview of endocannabinoid formation, deactivation, and biotransformation and outline the properties and therapeutic potential of pharmacological agents that interfere with those processes. We describe small-molecule inhibitors that target endocannabinoid-producing enzymes, carrier proteins that transport the endocannabinoids into cells, and intracellular endocannabinoid-metabolizing enzymes. We briefly discuss selected agents that simultaneous-ly interfere with components of the endocannabinoid system and with other functionally related signaling pathways.

The Endocannabinoid System in Caenorhabditis elegans.

The existence of a formal Endocannabinoid System in C. elegans has been questioned due to data showing the absence of typical cannabinoid receptors in the worm; however, the presence of a full metabolism for endocannabinoids, alternative ligands, and receptors for these agents and a considerable number of orthologous and homologous genes regulating physiological cannabinoid-like signals and responses - several of which are similar to those of mammals - demonstrates a well-structured and functional complex system in nematodes. In this review, we describe and compare similarities and differences between the Endocannabinoid System in mammals and nematodes, highlighting the basis for the integral study of this novel system in the worm.

GPR55 inhibits the pro-adipogenic activity of anandamide in human adipose stromal cells.

The endocannabinoid anandamide (AEA) stimulates adipogenesis via the cannabinoid receptor CB1 in adipose stromal cells (ASCs). However, AEA interacts also with nonclassical cannabinoid receptors, including transient receptor potential cation channel (TRPV)1 and G protein-coupled receptor (GPR)55. Their roles in AEA mediated adipogenesis of human ASCs have not been investigated. We examined the receptor-expressions by immunostaining on human ASCs and tested their functionality by measuring the expression of immediate early genes (IEGs) related to the transcription factor-complex AP-1 upon exposition to receptor agonists. Cells were stimulated with increasing concentrations of specific ligands to investigate the effects on ASC viability (proliferation and metabolic activity), secretory activity, and AEA mediated differentiation. ASCs expressed both receptors, and their activation suppressed IEG expression. TRPV1 did not affect viability or cytokine secretion. GPR55 decreased proliferation, and it inhibited the release of hepatocyte growth factor. Blocking GPR55 increased the pro-adipogenic activity of AEA. These data suggest that GPR55 functions as negative regulator of cannabinoid mediated pro-adipogenic capacity in ASCs.

Cannabinoid receptor 2 as a regulator of inflammation induced oleoylethanolamide in eosinophilic asthma.

BACKGROUND: Oleoylethanolamide (OEA), an endogenously generated cannabinoid-like compound, has been reported to be increased in patients with severe asthma and aspirin-exacerbated respiratory disease. Recruitment of activated eosinophils in the airways is a hallmark of bronchial asthma. OBJECTIVE: We explored the direct contribution of cannabinoid receptor 2 (CB2), a cognate receptor of OEA, which induces eosinophil activation in vitro and in vivo. METHODS: We investigated OEA signaling in the eosinophilic cell line dEol-1 in peripheral blood eosinophils from people with asthma. In order to confirm whether eosinophil activation by OEA is CB2 dependent or not, CB2 small interfering RNA and the CB2 antagonist SR144528 were used. The numbers of airway inflammatory cells and the levels of cytokines were measured in bronchoalveolar lavage fluid, and airway hyperresponsiveness was examined in the BALB/c mice. RESULTS: CB2 expression was increased after OEA treatment in both peripheral blood eosinophils and dEol-1 cells. It was also elevated after OEA-induced recruitment of eosinophils to the lungs in vivo. However, SR144528 treatment reduced the activation of peripheral blood eosinophils from asthmatic patients. Furthermore, CB2 knockdown decreased the activation of dEol-1 cells and the levels of inflammatory and type 2 cytokines. SR144528 treatment alleviated airway hyperresponsiveness and eosinophil recruitment to the lungs in vivo. CONCLUSION: CB2 may contribute to the pathogenesis of eosinophilic asthma. Our results provide new insight into the molecular mechanism of signal transduction by OEA in eosinophilic asthma.

Monoacylglycerol Lipase Protects the Presynaptic Cannabinoid 1 Receptor from Desensitization by Endocannabinoids after Persistent Inflammation.

Cannabinoid-targeted pain therapies are increasing with the expansion of cannabis legalization, however, their efficacy may be limited by pain-induced adaptations in the cannabinoid system. Cannabinoid receptor subtype 1 (CB1R) inhibition of spontaneous, GABAergic miniature IPSCs (mIPSCs) and evoked IPSCs (eIPSCs) in the ventrolateral periaqueductal gray (vlPAG) were compared in slices from naive and inflamed male and female Sprague Dawley rats. Complete Freund's Adjuvant (CFA) injections into the hindpaw induced persistent inflammation. In naive rats, exogenous cannabinoid agonists robustly reduce both eIPSCs and mIPSCs. After 5-7 d of inflammation, the effects of exogenous cannabinoids are significantly reduced because of CB1R desensitization via GRK2/3, as function is recovered in the presence of the GRK2/3 inhibitor, Compound 101 (Cmp101). Inhibition of GABA release by presynaptic µ-opioid receptors in the vlPAG does not desensitize with persistent inflammation. Unexpectedly, while CB1R desensitization significantly reduces the inhibition produced by exogenous agonists, depolarization-induced suppression of inhibition protocols that promote 2-arachidonoylglycerol (2-AG) synthesis exhibit prolonged CB1R activation after inflammation. 2-AG tone is detected in slices from CFA-treated rats when GRK2/3 is blocked, suggesting an increase in 2-AG synthesis after persistent inflammation. Inhibiting 2-AG degradation with the monoacylglycerol lipase (MAGL) inhibitor JZL184 during inflammation results in the desensitization of CB1Rs by endocannabinoids that is reversed with Cmp101. Collectively, these data indicate that persistent inflammation primes CB1Rs for desensitization, and MAGL degradation of 2-AG protects CB1Rs from desensitization in inflamed rats. These adaptations with inflammation have important implications for the development of cannabinoid-based pain therapeutics targeting MAGL and CB1Rs.SIGNIFICANCE STATEMENT Presynaptic G-protein-coupled receptors are resistant to desensitization. Here we find that persistent inflammation increases endocannabinoid levels, priming presynaptic cannabinoid 1 receptors for desensitization on subsequent addition of exogenous agonists. Despite the reduced efficacy of exogenous agonists, endocannabinoids have prolonged efficacy after persistent inflammation. Endocannabinoids readily induce cannabinoid 1 receptor desensitization if their degradation is blocked, indicating that endocannabinoid concentrations are maintained at subdesensitizing levels and that degradation is critical for maintaining endocannabinoid regulation of presynaptic GABA release in the ventrolateral periaqueductal gray during inflammatory states. These adaptations with inflammation have important implications for the development of cannabinoid-based pain therapies.

Cannabidiol protects C2C12 myotubes against cisplatin-induced atrophy by regulating oxidative stress.

Cancer and chemotherapy induce a severe loss of muscle mass (known as cachexia), which negatively impact cancer treatment and patient survival. The aim of the present study was to investigate whether cannabidiol (CBD) administration may potentially antagonize the effects of cisplatin in inducing muscle atrophy, using a model of myotubes in culture. Cisplatin treatment resulted in a reduction of myotube diameter (15.7 ± 0.3 vs. 22.2 ± 0.5 µm, P < 0.01) that was restored to control level with 5 µM CBD (20.1 ± 0.4 µM, P < 0.01). Protein homeostasis was severely altered with a ≈70% reduction in protein synthesis (P < 0.01) and a twofold increase in proteolysis (P < 0.05) in response to cisplatin. Both parameters were dose dependently restored by CBD cotreatment. Cisplatin treatment was associated with increased thiobarbituric acid reactive substances (TBARS) content (0.21 ± 0.03 to 0.48 ± 0.03 nmol/mg prot, P < 0.05), catalase activity (0.24 ± 0.01 vs. 0.13 ± 0.02 nmol/min/µg prot, P < 0.01), whereas CBD cotreatment normalized TBARS content to control values (0.22 ± 0.01 nmol/mg prot, P < 0.01) and reduced catalase activity (0.17 ± 0.01 nmol/min/µg prot, P < 0.05). These changes were associated with increased mRNA expression of GPX1, SOD1, SOD2, and CAT mRNA expression in response to cisplatin (P < 0.01), which was corrected by CBD cotreatment (P < 0.05). Finally, cisplatin treatment increased the mitochondrial protein content of NDUFB8, UQCRC2, COX4, and VDAC1 (involved in mitochondrial respiration and apoptosis), and CBD cotreatment restored their expression to control values. Altogether, our results demonstrated that CBD antagonize the cisplatin-induced C2C12 myotube atrophy and could be used as an adjuvant in the treatment of cancer cachexia to help maintain muscle mass and improve patient quality of life.NEW & NOTEWORTHY In an in vitro model, cisplatin treatment led to myotube atrophy associated with dysregulation of protein homeostasis and increased oxidative stress, resulting in increased apoptosis. Cotreatment with cannabidiol was able to prevent this phenotype by promoting protein homeostasis and reducing oxidative stress.

Pharmaceutical targeting of the cannabinoid type 1 receptor impacts the crosstalk between immune cells and islets to reduce insulitis in humans.

AIMS/HYPOTHESIS: Insulitis, a hallmark of inflammation preceding autoimmune type 1 diabetes, leads to the eventual loss of functional beta cells. However, functional beta cells can persist even in the face of continuous insulitis. Despite advances in immunosuppressive treatments, maintaining functional beta cells to prevent insulitis progression and hyperglycaemia remains a challenge. The cannabinoid type 1 receptor (CB1R), present in immune cells and beta cells, regulates inflammation and beta cell function. Here, we pioneer an ex vivo model mirroring human insulitis to investigate the role of CB1R in this process. METHODS: CD4+ T lymphocytes were isolated from peripheral blood mononuclear cells (PBMCs) from male and female individuals at the onset of type 1 diabetes and from non-diabetic individuals, RNA was extracted and mRNA expression was analysed by real-time PCR. Single beta cell expression from donors with type 1 diabetes was obtained from data mining. Patient-derived human islets from male and female cadaveric donors were 3D-cultured in solubilised extracellular matrix gel in co-culture with the same donor PBMCs, and incubated with cytokines (IL-1ß, TNF-α, IFN-γ) for 24-48 h in the presence of vehicle or increasing concentrations of the CB1R blocker JD-5037. Expression of CNR1 (encoding for CB1R) was ablated using CRISPR/Cas9 technology. Viability, intracellular stress and signalling were assayed by live-cell probing and real-time PCR. The islet function measured as glucose-stimulated insulin secretion was determined in a perifusion system. Infiltration of immune cells into the islets was monitored by microscopy. Non-obese diabetic mice aged 7 weeks were treated for 1 week with JD-5037, then euthanised. Profiling of immune cells infiltrated in the islets was performed by flow cytometry. RESULTS: CNR1 expression was upregulated in circulating CD4+ T cells from individuals at type 1 diabetes onset (6.9-fold higher vs healthy individuals) and in sorted islet beta cells from donors with type 1 diabetes (3.6-fold higher vs healthy counterparts). The peripherally restricted CB1R inverse agonist JD-5037 arrested the initiation of insulitis in humans and mice. Mechanistically, CB1R blockade prevented islet NO production and ameliorated the ATF6 arm of the unfolded protein response. Consequently, cyto/chemokine expression decreased in human islets, leading to sustained islet cell viability and function. CONCLUSIONS/INTERPRETATION: These results suggest that CB1R could be an interesting target for type 1 diabetes while highlighting the regulatory mechanisms of insulitis. Moreover, these findings may apply to type 2 diabetes where islet inflammation is also a pathophysiological factor. DATA AVAILABILITY: Transcriptomic analysis of sorted human beta cells are from Gene Expression Omnibus database, accession no. GSE121863, available at https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSM3448161 .

Role of omega-3 endocannabinoids in the modulation of T-cell activity in a multiple sclerosis experimental autoimmune encephalomyelitis (EAE) model.

Epidemiological studies show that omega-3 fatty acid consumption is associated with improved conditions in neurodegenerative diseases such as multiple sclerosis (MS). However, the mechanism of this association is not well understood. Emerging evidence suggests that parent molecules such as docosahexaenoic acid are converted into downstream metabolites that are capable of directly modulating immune responses. In vitro, we found that docosahexaenoyl ethanolamide (DHEA), another dietary component and its epoxide metabolite, reduced the polarization of naïve T-cells toward proinflammatory Th1 and Th17 phenotypes. Furthermore, we identified that DHEA and related endocannabinoids are changing during the disease progression in mice undergoing relapse-remitting experimental autoimmune encephalomyelitis (RR-EAE). In addition, daily administration of DHEA to mice delayed the onset of disease, the rate of relapse, and the severity of clinical scores at relapse in RR-EAE, an animal model of MS. Collectively, these data indicate that DHEA and their downstream metabolites reduce the disease severity in the RR-EAE model of MS and can be potential dietary adjuvants to existing MS therapeutics.

Exploring the versatile roles of the endocannabinoid system and phytocannabinoids in modulating bacterial infections.

The endocannabinoid system (ECS), initially identified for its role in maintaining homeostasis, particularly in regulating brain function, has evolved into a complex orchestrator influencing various physiological processes beyond its original association with the nervous system. Notably, an expanding body of evidence emphasizes the ECS's crucial involvement in regulating immune responses. While the specific role of the ECS in bacterial infections remains under ongoing investigation, compelling indications suggest its active participation in host-pathogen interactions. Incorporating the ECS into the framework of bacterial pathogen infections introduces a layer of complexity to our understanding of its functions. While some studies propose the potential of cannabinoids to modulate bacterial function and immune responses, the outcomes inherently hinge on the specific infection and cannabinoid under consideration. Moreover, the bidirectional relationship between the ECS and the gut microbiota underscores the intricate interplay among diverse physiological processes. The ECS extends its influence far beyond its initial discovery, emerging as a promising therapeutic target across a spectrum of medical conditions, encompassing bacterial infections, dysbiosis, and sepsis. This review comprehensively explores the complex roles of the ECS in the modulation of bacteria, the host's response to bacterial infections, and the dynamics of the microbiome. Special emphasis is placed on the roles of cannabinoid receptor types 1 and 2, whose signaling intricately influences immune cell function in microbe-host interactions.

Mu-opioid receptor activation in the habenula modulates synaptic transmission and depression-like behaviors.

Opioid system dysregulation in response to stress is known to lead to psychiatric disorders including major depression. Among three different types of opioid receptors, the mu-type receptors (mORs) are highly expressed in the habenula complex, however, the action of mORs in this area and its interaction with stress exposure is largely unknown. Therefore, we investigated the roles of mORs in the habenula using male rats of an acute learned helplessness (aLH) model. First, we found that mOR activation decreased both excitatory and inhibitory synaptic transmission onto the lateral habenula (LHb). Intriguingly, this mOR-induced synaptic depression was reduced in an animal model of depression compared to that of controls. In naïve animals, we found an unexpected interaction between mORs and the endocannabinoid (eCB) signaling occurring in the LHb, which mediates presynaptic alteration occurring with mOR activation. However, we did not observe presynaptic alteration by mOR activation after stress exposure. Moreover, selective mOR activation in the habenula before, but not after, stress exposure effectively reduced helpless behaviors compared to aLH animals. Our observations are consistent with clinical reports suggesting the involvement of mOR signaling in depression, and additionally reveal a critical time window of mOR action in the habenula for ameliorating helplessness symptoms.

Modulation of Oxidative Stress and Neuroinflammation by Cannabidiol (CBD): Promising Targets for the Treatment of Alzheimer's Disease.

Alzheimer's disease (AD) is a progressive neurodegenerative disease and the most common form of dementia globally. Although the direct cause of AD remains under debate, neuroinflammation and oxidative stress are critical components in its pathogenesis and progression. As a result, compounds like cannabidiol (CBD) are being increasingly investigated for their ability to provide antioxidant and anti-inflammatory neuroprotection. CBD is the primary non-psychotropic phytocannabinoid derived from Cannabis sativa. It has been found to provide beneficial outcomes in a variety of medical conditions and is gaining increasing attention for its potential therapeutic application in AD. CBD is not psychoactive and its lipophilic nature allows its rapid distribution throughout the body, including across the blood-brain barrier (BBB). CBD also possesses anti-inflammatory, antioxidant, and neuroprotective properties, making it a viable candidate for AD treatment. This review outlines CBD's mechanism of action, the role of oxidative stress and neuroinflammation in AD, and the effectiveness and limitations of CBD in preclinical models of AD.

Cannabidiol attenuates seizure susceptibility and behavioural deficits in adult CDKL5R59X knock-in mice.

Cyclin-dependent kinase-like 5 (CDKL5) deficiency disorder (CDD) is caused by a loss-of-function mutation in CDKL5 gene, encoding a serine-threonine kinase highly expressed in the brain. CDD manifests with early-onset epilepsy, autism, motor impairment and severe intellectual disability. While there are no known treatments for CDD, the use of cannabidiol has recently been introduced into clinical practice for neurodevelopmental disorders. Given the increased clinical utilization of cannabidiol, we examined its efficacy in the CDKL5R59X knock-in (R59X) mice, a CDD model based on a human mutation that exhibits both lifelong seizure susceptibility and behavioural deficits. We found that cannabidiol pre-treatment rescued the increased seizure susceptibility in response to the chemoconvulsant pentylenetetrazol (PTZ), attenuated working memory and long-term memory impairments, and rescued social deficits in adult R59X mice. To elucidate a potential mechanism, we compared the developmental hippocampal and cortical expression of common endocannabinoid (eCB) targets in R59X mice and their wild-type littermates, including cannabinoid type 1 receptor (CB1R), transient receptor potential vanilloid type 1 (TRPV1) and 2 (TRPV2), G-coupled protein receptor 55 (GPR55) and adenosine receptor 1 (A1R). Many of these eCB targets were developmentally regulated in both R59X and wild-type mice. In addition, adult R59X mice demonstrated significantly decreased expression of CB1R and TRPV1 in the hippocampus, and TRPV2 in the cortex, while TRPV1 was increased in the cortex. These findings support the potential for dysregulation of eCB signalling as a plausible mechanism and therapeutic target in CDD, given the efficacy of cannabidiol to attenuate hyperexcitability and behavioural deficits in this disorder.

Angiotensinergic effect of ß-Caryophyllene on Lipopolysaccharide- induced systemic inflammation.

Renin-Angiotensin System (RAS) is a peptidergic system, canonically known for its role in blood pressure regulation. Furthermore, a non-canonical RAS regulates pathophysiological phenomena, such as inflammation since it consists of two main axes: the pro-inflammatory renin/(pro)renin receptor ((P)RR) axis, and the anti-inflammatory angiotensin-converting enzyme 2 (ACE2)/Angiotensin-(1-7) (Ang-(1-7))/Mas Receptor (MasR) axis. Few phytochemicals have shown to exert angiotensinergic and anti-inflammatory effects through some of these axes; nevertheless, anti-inflammatory drugs, such as phytocannabinoids have not been studied regarding this subject. Among phytocannabinoids, ß-Caryophyllene stands out as a dietary phytocannabinoid with antiphlogistic activity that possess a unique sesquiterpenoid structure. Although its cannabinergic effect has been studied, its angiotensinergic effect reminds underexplored. This study aims to explore the angiotensinergic effect of ß-Caryophyllene on inflammation and stress at a systemic level. After intranasal Lipopolysaccharide (LPS) installation and oral treatment with ß-Caryophyllene, the concentration and activity of key RAS elements in the serum, such as Renin, ACE2 and Ang-(1-7), along with the stress hormone corticosterone and pro/anti-inflammatory cytokines, were measured in mice serum. The results show that ß-Caryophyllene treatment modified RAS levels by increasing Renin and Ang-(1-7), alongside the reduction of pro-inflammatory cytokines and corticosterone levels. These results indicate that ß-Caryophyllene exhibits angiotensinergic activity in favor of anti-inflammation.

Probing molecular pathways: Illuminating the connection between COVID-19 and Alzheimer's disease through the endocannabinoid system dynamics.

Our study investigates the molecular link between COVID-19 and Alzheimer's disease (AD). We aim to elucidate the mechanisms by which COVID-19 may influence the onset or progression of AD. Using bioinformatic tools, we analyzed gene expression datasets from the Gene Expression Omnibus (GEO) database, including GSE147507, GSE12685, and GSE26927. Intersection analysis was utilized to identify common differentially expressed genes (CDEGs) and their shared biological pathways. Consensus clustering was conducted to group AD patients based on gene expression, followed by an analysis of the immune microenvironment and variations in shared pathway activities between clusters. Additionally, we identified transcription factor-binding sites shared by CDEGs and genes in the common pathway. The activity of the pathway and the expression levels of the CDEGs were validated using GSE164805 and GSE48350 datasets. Six CDEGs (MAL2, NECAB1, SH3GL2, EPB41L3, MEF2C, and NRGN) were identified, along with a downregulated pathway, the endocannabinoid (ECS) signaling pathway, common to both AD and COVID-19. These CDEGs showed a significant correlation with ECS activity (p < 0.05) and immune functions. The ECS pathway was enriched in healthy individuals' brains and downregulated in AD patients. Validation using GSE164805 and GSE48350 datasets confirmed the differential expression of these genes in COVID-19 and AD tissues. Our findings reveal a potential pathogenetic link between COVID-19 and AD, mediated by CDEGs and the ECS pathway. However, further research and multicenter evidence are needed to translate these findings into clinical applications.

Quantal size increase induced by the endocannabinoid 2-arachidonoylglycerol requires activation of CGRP receptors in mouse motor synapses.

In mouse motor synapses, the exogenous application of the endocannabinoid (EC) 2-arachidonoylglycerol (2-AG) increases acetylcholine (ACh) quantal size due to the activation of CB1 receptors and the stimulation of ACh vesicular uptake. In the present study, microelectrode recordings of miniature endplate potentials (MEPP) revealed that this effect of 2-AG is independent of brain-derived neurotrophic factor (BDNF) signaling but involves the activation of calcitonin gene-related peptide (CGRP) receptors along with CB1 receptors. Potentiation of MEPP amplitude in the presence of 2-AG was prevented by blockers of CGRP receptors and ryanodine receptors (RyR) and by inhibitors of phospholipase C (PLC) and Ca2+ /calmodulin-dependent protein kinase II (CaMKII). Therefore, we suggest a hypothetical chain of events, which starts from the activation of presynaptic CB1 receptors, involves PLC, RyR, and CaMKII, and results in CGRP release with the subsequent activation of presynaptic CGRP receptors. Activation of CGRP receptors is probably a part of a complex molecular cascade leading to the 2-AG-induced increase in ACh quantal size and MEPP amplitude. We propose that the same chain of events may also take place if 2-AG is endogenously produced in mouse motor synapses, because the increase in MEPP amplitude that follows after prolonged tetanic muscle contractions (30 Hz, 2 min) was prevented by the blocking of CB1 receptors. This work may help to unveil the previously unknown aspects of the functional interaction between ECs and peptide modulators aimed at the regulation of quantal size and synaptic transmission.