Cannabinoid Receptor Research Articles

Crystal structure, Hirshfeld surface analysis, computational study and molecular docking simulation of 4-aminoantipyrine derivative

Herein, a new derivative (4APSA) has been synthesized by treating 4-aminoantipyrine with succinic anhydride at 25 °C employing glacial acetic acid as a solvent. The molecular structure of 4APSA is investigated via single crystal X-ray diffraction (XRD) analysis. Cambridge structure database CSD 2023.3.1 (updated March 2024) was used for comparison of 4APSA with the associated reported structures. The supramolecular assemblage was alleviated by several intermolecular connections. Hirshfeld surface analysis was conducted for further assessment of intermolecular interactions with respect to interatomic contacts for 4APSA and a closely associated structure having a reference code of HOMDUI. Interaction energy calculations were performed at B3LYP/6–31G(d,p) electron density level to find the type of interaction energy that had the highest contribution to the supramolecular assembly. Moreover, in-depth DFT calculations were studied at B3LYP/6-311++g(d,p) level to obtain molecular structure, Mulliken charge analysis, MEP surface, electronic properties, quantum chemical reactivity, and DOS spectrum. Strong correlations are found between theoretical and experimental structures. The ADME-T prediction was applied for the 4APSA, 4AP, and SA compounds utilizing OSIRIS DataWarrior. Docking simulation demonstrates the high tendency of 4APSA to bind with the active sites of Cannabinoid Receptor Subtype 2 (CB2R) compared to JWH-018. This finding suggested that the synthesized compound has a positive effect on the CB2R protein.

Actomyosin-mediated inhibition of synaptic vesicle release under CB1R activation

Long-term synaptic plasticity is critical for adaptive function of the brain, but presynaptic mechanisms of functional plasticity remain poorly understood. Here, we show that changes in synaptic efficacy induced by activation of the cannabinoid type-1 receptor (CB1R), one of the most widespread G-protein coupled receptors in the brain, requires contractility of the neuronal actomyosin cytoskeleton. Specifically, using a synaptophysin-pHluorin probe (sypH2), we show that inhibitors of non-muscle myosin II (NMII) ATPase as well as one of its upstream effectors Rho-associated kinase (ROCK) prevent the reduction of synaptic vesicle release induced by CB1R activation. Using 3D STORM super-resolution microscopy, we find that activation of CB1R induces a redistribution of synaptic vesicles within presynaptic boutons in an actomyosin dependent manner, leading to vesicle clustering within the bouton and depletion of synaptic vesicles from the active zone. We further show, using sypH2, that inhibitors of NMII and ROCK specifically restore the release of the readily releasable pool of synaptic vesicles from the inhibition induced by CB1R activation. Finally, using slice electrophysiology, we find that activation of both NMII and ROCK is necessary for the long-term, but not the short-term, form of CB1R induced synaptic plasticity at excitatory cortico-striatal synapses. We thus propose a novel mechanism underlying CB1R-induced plasticity, whereby CB1R activation leads to a contraction of the actomyosin cytoskeleton inducing a reorganization of the functional presynaptic vesicle pool, preventing vesicle release and inducing long-term depression.

Enhancing Drug Discovery and Development through the Integration of Medicinal Chemistry, Chemical Biology, and Academia-Industry Partnerships: Insights from Roche's Endocannabinoid System Projects.

The endocannabinoid system (ECS) is a critical regulatory network composed of endogenous cannabinoids (eCBs), their synthesizing and degrading enzymes, and associated receptors. It is integral to maintaining homeostasis and orchestrating key functions within the central nervous and immune systems. Given its therapeutic significance, we have launched a series of drug discovery endeavors aimed at ECS targets, including peroxisome proliferator-activated receptors (PPARs), cannabinoid receptors types 1 (CB1R) and 2 (CB2R), and monoacylglycerol lipase (MAGL), addressing a wide array of medical needs. The pursuit of new therapeutic agents has been enhanced by the creation of specialized labeled chemical probes, which aid in target localization, mechanistic studies, assay development, and the establishment of biomarkers for target engagement. By fusing medicinal chemistry with chemical biology in a comprehensive, translational end-to-end drug discovery strategy, we have expedited the development of novel therapeutics. Additionally, this strategy promises to foster highly productive partnerships between industry and academia, as will be illustrated through various examples.

Recent Molecular Targets and Their Ligands for the Treatment of Alzheimer Disease.

Alzheimer's disease is a multifaceted neurodegenerative disease. Cholinergic dysfunction, amyloid β toxicity, tauopathies, oxidative stress, neuroinflammation are among the main pathologies of the disease. Ligands targeting more than one pathology, multi-target directed ligands, attract attention in the recent years to tackle Alzheimer's disease. In this review, we aimed to cover different biochemical pathways, that are revealed in recent years for the pathology of the disease, as druggable targets such as cannabinoid receptors, matrix metalloproteinases, histone deacetylase and various kinases including, glycogen synthase kinase-3, mitogen-activated protein kinase and c-Jun N-terminal kinase, and their ligands for the treatment of Alzheimer's disease in the hope of providing more realistic insights into the field.

Cannabinoid CB2 receptors enhance high-fat diet evoked peripheral neuroinflammation

It is known that the cannabinoid type 2 (CB2) receptor has an anti-inflammatory role. Therefore, animals without CB2 receptors show enhanced inflammation and pain in the model of chronic pain, e.g., neuropathic pain. We previously proposed the upregulated leptin signaling at the peripheral nerve as one of the underlying molecular mechanisms of pain exacerbation in nerve-injured CB2 knockouts, as they displayed robust upregulation of leptin receptors and leptin signaling in the peripheral nerve. Due to these past results, we hypothesized that CB2 receptor deficiency might also modify the peripheral neuroinflammation led by chronic exposure to a high-fat diet (HFD). Interestingly, CB2 knockout animals showed significant resistance to HFD-induced neuroinflammation. Namely, 5-week feeding of HFD induced substantial hypersensitivity in WT animals, while tactile sensitivity of HFD-fed CB2 knockouts remained intact. HFD-fed WT animals also displayed the robust upregulation of chemokine CXCR4 expression with increased macrophage infiltration, which was never observed in HFD-fed CB2 knockout mice. Moreover, 5-week HFD exposure led significant increase of CD11b+Ly6G−Ly6Chigh cells and a decrease of CD11b+Ly6G+Ly6Clow cells in the spleen of WT animals, which was also not found in either HFD-fed CB2 knockouts or standard diet-fed WT and CB2 animals. Together with past reports, these results suggest that CB2 receptors might have a double-sided regulatory role in the context of inflammation development or, more widely, immune system regulation. We propose that CB2 signaling is not always anti-inflammatory and could take a pro-inflammatory role depending on the cause of the inflammation.

Region-Specific Gene Expression Changes Associated with Oleoylethanolamide-Induced Attenuation of Alcohol Self-Administration.

Oleoylethanolamide (OEA) is a lipid with anti-inflammatory activity that modulates multiple reward-related behaviors. Previous studies have shown that OEA treatment reduces alcohol self-administration (SA) while inhibiting alcohol-induced inflammatory signaling. Nevertheless, the specific mechanisms that OEA targets to achieve these effects have not been widely explored. Here, we tested the effects of OEA treatment during alcohol SA, extinction or previous to cue-induced reinstatement of alcohol seeking. In addition, we measured gene expression changes in the striatum and hippocampus of relevant receptors for alcohol consumption (Drd1, Drd2, Cnr1, Oprm) as well as immune-related proteins (Il-6, Il-1β, Tlr4) and the brain-derived neurotrophic factor (Bdnf). Our results confirmed that when administered contingently, systemic OEA administration reduced alcohol SA and attenuated cue-induced reinstatement. Interestingly, we also observed that OEA treatment reduced the number of sessions needed for the extinction of alcohol seeking. Biochemical analyses showed that OEA induced gene expression changes in dopamine and cannabinoid receptors in the striatum and hippocampus. In addition, OEA treatment modulated the long-term immune response and increased Bdnf expression. These results suggest that boosting OEA levels may be an effective strategy for reducing alcohol SA and preventing relapse.

CB2 expression in mouse brain: from mapping to regulation in microglia under inflammatory conditions

Since its detection in the brain, the cannabinoid receptor type 2 (CB2) has been considered a promising therapeutic target for various neurological and psychiatric disorders. However, precise brain mapping of its expression is still lacking. Using magnetic cell sorting, calibrated RT-qPCR and single-nucleus RNAseq, we show that CB2 is expressed at a low level in all brain regions studied, mainly by few microglial cells, and by neurons in an even lower proportion. Upon lipopolysaccharide stimulation, modeling neuroinflammation in non-sterile conditions, we demonstrate that the inflammatory response is associated with a transient reduction in CB2 mRNA levels in brain tissue, particularly in microglial cells. This result, confirmed in the BV2 microglial cell line, contrasts with the positive correlation observed between CB2 mRNA levels and the inflammatory response upon stimulation by interferon-gamma, modeling neuroinflammation in sterile condition. Discrete brain CB2 expression might thus be up- or down-regulated depending on the inflammatory context.

Basal forebrain-lateral habenula inputs and control of impulsive behavior.

Deficits in impulse control are observed in several neurocognitive disorders, including attention deficit hyperactivity (ADHD), substance use disorders (SUDs), and those following traumatic brain injury (TBI). Understanding brain circuits and mechanisms contributing to impulsive behavior may aid in identifying therapeutic interventions. We previously reported that intact lateral habenula (LHb) function is necessary to limit impulsivity defined by impaired response inhibition in rats. Here, we examine the involvement of a synaptic input to the LHb on response inhibition using cellular, circuit, and behavioral approaches. Retrograde fluorogold tracing identified basal forebrain (BF) inputs to LHb, primarily arising from ventral pallidum and nucleus accumbens shell (VP/NAcs). Glutamic acid decarboxylase and cannabinoid CB1 receptor (CB1R) mRNAs colocalized with fluorogold, suggesting a cannabinoid modulated GABAergic pathway. Optogenetic activation of these axons strongly inhibited LHb neuron action potentials and GABA release was tonically suppressed by an endogenous cannabinoid in vitro. Behavioral experiments showed that response inhibition during signaled reward omission was impaired when VP/NAcs inputs to LHb were optogenetically stimulated, whereas inhibition of this pathway did not alter LHb control of impulsivity. Systemic injection with the psychotropic phytocannabinoid, Δ9-tetrahydrocannabinol (Δ9-THC), also increased impulsivity in male, and not female rats, and this was blocked by LHb CB1R antagonism. However, as optogenetic VP/NAcs pathway inhibition did not alter impulse control, we conclude that the pro-impulsive effects of Δ9-THC likely do not occur via inhibition of this afferent. These results identify an inhibitory LHb afferent that is controlled by CB1Rs that can regulate impulsive behavior.

Bioelectronic sensing platform emulating the human endocannabinoid system for assessing and modulating of cannabinoid activity

Cannabinoids are involved in physiological and neuromodulatory processes through their interactions with the human cannabinoid receptor-based endocannabinoid system. Their association with neurodegenerative diseases and brain reward pathways underscores the importance of evaluating and modulating cannabinoid activity for both understanding physiological mechanisms and developing therapeutic drugs. The use of agonists and antagonists could be strategic approaches for modulation. In this study, we introduce a bioelectronic sensor designed to monitor cannabinoid binding to receptors and assess their agonistic and antagonistic properties. We produced human cannabinoid receptor 1 (hCB1R) via an Escherichia coli expression system and incorporated it into nanodiscs (NDs). These hCB1R-NDs were then immobilized on a single-walled carbon nanotube field-effect transistor (swCNT-FET) to construct a bioelectronic sensing platform. This novel system can sensitively detect the cannabinoid ligand anandamide (AEA) at concentrations as low as 1 fM, demonstrating high selectivity and real-time response. It also successfully identified the hCB1R agonist Δ9-tetrahydrocannabinol and observed that the hCB1R antagonist rimonabant diminished the sensor signal upon AEA binding, indicating the antagonism-based modulation of ligand interaction. Consequently, our bioelectronic sensing platform holds potential for ligand detection and analysis of agonism and antagonism.

β-Caryophyllene mitigates ischemic stroke-induced white matter lesions by inhibiting pyroptosis

β-Caryophyllene (BCP), a selective agonist for cannabinoid receptor 2 (CB2R), has demonstrated promising protective effects in various pathological conditions. However, the neuroprotective effects of BCP on white matter damage induced by ischemic stroke have not been elucidated previously. In this study, we find that BCP not only improves sensorimotor and cognitive function via CB2R but also mitigates white matter lesions in mice following ischemic stroke. Furthermore, BCP enhances the viability of MO3.13 oligodendrocytes after oxygen-glucose deprivation and reoxygenation (OGD/R), attenuating OGD/R-induced cellular damage and pyroptosis. Notably, these protective effects of BCP are partially enhanced by the NLRP3 inhibitor MCC950 and counteracted by the NLRP3 activator nigericin. In addition, nigericin significantly exacerbates neurological outcomes and increases white matter lesions following BCP treatment in middle cerebral artery occlusion (MCAO) mice. These results suggest that BCP may ameliorate neurological deficits and white matter damage induced by cerebral ischemia through inhibiting NLRP3-mediated pyroptosis.

Cannabinoid receptor 2 agonist AM1241 alleviates epileptic seizures and epilepsy-associated depression via inhibiting neuroinflammation in a pilocarpine-induced chronic epilepsy mouse model

Increasing evidence suggests that cannabinoid receptor 2 (CB2R) serves as a promising anti-inflammatory target. While inflammation is known to play crucial roles in the pathogenesis of epilepsy, the involvement of CB2R in epilepsy remains unclear. This study aimed to investigate the effects of a CB2R agonist, AM1241, on epileptic seizures and depressive-like behaviors in a mouse model of chronic epilepsy induced by pilocarpine. A chronic epilepsy mouse model was established by intraperitoneal administration of pilocarpine. The endogenous cannabinoid system (eCBs) in the hippocampus was examined after status epilepticus (SE). Animals were then treated with AM1241 and compared with a vehicle-treated control group. Additionally, the role of the AMPK/NLRP3 signaling pathway was explored using the selective AMPK inhibitor dorsomorphin. Following SE, CB2R expression increased significantly in hippocampal microglia. Administration of AM1241 significantly reduced seizure frequency, immobility time in the tail suspension test, and neuronal loss in the hippocampus. In addition, AM1241 treatment attenuated microglial activation, inhibited pro-inflammatory polarization of microglia, and suppressed NLRP3 inflammasome activation in the hippocampus after SE. Further, the therapeutic effects of AM1241 were abolished by the AMPK inhibitor dorsomorphin. Our findings suggest that CB2R agonist AM1241 may alleviate epileptic seizures and its associated depression by inhibiting neuroinflammation through the AMPK/NLRP3 signaling pathway. These results provide insight into a novel therapeutic approach for epilepsy.

Dietary limonene promotes gastrointestinal barrier function via upregulating tight/adherens junction proteins through cannabinoid receptor type-1 antagonistic mechanism and alters cellular metabolism in intestinal epithelial cells.

Limonene, a dietary monocyclic monoterpene commonly found in citrus fruits and various aromatic plants, has garnered increasing interest as a gastrointestinal protectant. This study aimed to assess the effects of limonene on intestinal epithelial barrier function and investigate the involvement of cannabinoid receptor type-1 (CB1R) in vitro. Additionally, the study focused on examining the metabolomic changes induced by limonene in the intestinal epithelial cells (Caco-2). Initial analysis of transepithelial electrical resistance (TEER) revealed that both l-limonene and d-limonene, isomers of limonene, led to a dose- and time-dependent increase in TEER in normal cells and those inflamed by pro-inflammatory cytokines mixture (CytoMix). Furthermore, both types of limonene reduced CytoMix-induced paracellular permeability, as demonstrated by a decrease in Lucifer yellow flux. Moreover, d-limonene and l-limonene treatment increased the expression of tight junction molecules (TJs) such as occludin, claudin-1, and ZO-1, at both the transcriptional and translational levels. d-Limonene upregulates E-cadherin, a molecule involved in adherens junctions (AJs). Mechanistic investigations demonstrated that d-limonene and l-limonene treatment significantly inhibited CB1R at the protein, while the mRNA level remained unchanged. Notably, the inhibitory effect of d-limonene on CB1R was remarkably similar to that of pharmacological CB1R antagonists, such as rimonabant and ORG27569. d-limonene also alters Caco-2 cell metabolites. A substantial reduction in β-glucose and 2-succinamate was detected, suggesting limonene may impact intestinal epithelial cells' glucose uptake and glutamate metabolism. These findings suggest that d-limonene's CB1R antagonistic property could effectively aid in the recovery of intestinal barrier damage, marking it a promising gastrointestinal protectant.

Bidirectional Effect of Long-Term Δ9-Tetrahydrocannabinol Treatment on mTOR Activity and Metabolome.

Brain aging is associated with cognitive decline, reduced synaptic plasticity, and altered metabolism. The activity of mechanistic target of rapamycin (mTOR) has a major impact on aging by regulating cellular metabolism. Although reduced mTOR signaling has a general antiaging effect, it can negatively affect the aging brain by reducing synaptogenesis and thus cognitive functions. Increased mTOR activity facilitates aging and is responsible for the amnestic effect of the cannabinoid receptor 1 agonist Δ9-tetrahydrocannabinol (THC) in higher doses. Long-term low-dose Δ9-THC had an antiaging effect on the brain by restoring cognitive abilities and synapse densities in old mice. Whether changes in mTOR signaling and metabolome are associated with its positive effects on the aging brain is an open question. Here, we show that Δ9-THC treatment has a tissue-dependent and dual effect on mTOR signaling and the metabolome. In the brain, Δ9-THC treatment induced a transient increase in mTOR activity and in the levels of amino acids and metabolites involved in energy production, followed by an increased synthesis of synaptic proteins. Unexpectedly, we found a similar reduction in the mTOR activity in adipose tissue and in the level of amino acids and carbohydrate metabolites in blood plasma as in animals on a low-calorie diet. Thus, long-term Δ9-THC treatment first increases the level of energy and synaptic protein production in the brain, followed by a reduction in mTOR activity and metabolic processes in the periphery. Our study suggests that a dual effect on mTOR activity and the metabolome could be the basis for an effective antiaging and pro-cognitive medication.

Effect of EMB-FUBINACA on brain endothelial cell angiogenesis: Expression analysis of angiogenic markers.

Angiogenesis is the process by which blood vessels are generated from preexisting ones. Synthetic cannabinoids represent new psychoactive substances that bind to the cannabinoid receptor 1 (CB1R) and cannabinoid receptor 2 (CB2R) and simulate similar effects of tetrahydrocannabinol, the primary component found in cannabis. In the present study, we used the synthetic cannabinoid EMB-FUBINACA to study its impact on brain angiogenesis. Human brain microvascular endothelial cells (HBMECs) were cultivated in DMEM media before being subjected to different concentrations of EMB-FUBINACA and the control. Cell viability and the migration rates of HBMECs were evaluated using the viability and wound healing assays, respectively. An in vitro Matrigel Tube Formation Assay was carried out to measure the angiogenic capacity of endothelial cells. Angiopoietin-1 (ANG-1), Angiopoietin-2 (ANG-2), and vascular endothelial growth factor (VEGF) mRNA expression were detected using Real-Time PCR. The released VEGF, ANG-1, and ANG-2 concentrations were detected using ELISA. Western blotting was performed to measure the levels of phosphorylated GSK-3β, VEGF, ANG-1, and ANG-2. EMB-FUBINACA stimulated endothelial cell proliferation, migration, and capillary tube-like formation and promoted the expression of proangiogenic factors on RNA and protein levels. This study points out that the synthetic cannabinoid EMB-FUBINACA is a potential candidate for further investigations to confirm its potential as an inducer of brain angiogenesis. This could encourage researchers to create a new therapeutic approach for angiogenesis-related diseases.

Activation of CB2 Receptors by (-)-Cannabichromene but Not (+)-Cannabichromene.

Introduction: Cannabichromene (CBC) is a minor constituent of cannabis that is a selective cannabinoid CB2 receptor agonist and activator of TRPA1. To date, it has not been shown whether (-)-CBC, (+)-CBC, or both can mediate these effects. In this study, we investigate the activity of the CBC enantiomers at CB1, CB2, and Transient receptor potential ankyrin 1 (TRPA1) receptors in vitro. Materials and Methods: CBC enantiomers were purified from synthetic CBC by chiral chromatography, and their optical activity was confirmed by spectroscopy. Human CB1 and CB2 receptor activity was measured using a fluorescent assay of membrane potential in stably transfected AtT20 cells. TRPA1 activation was measured using a fluorescent assay of intracellular calcium in stably transfected HEK293 cells. Results: The (-)-CBC activated CB2 with an EC50 of 1.5 µM, to a maximum of 60% of (-)CP55940. (+)-CBC did not activate CB2 at concentrations up to 30 µM. Only 30 µM (-)-CBC produced detectable activation of CB1, (+)-CBC was inactive. Both (-)-CBC and (+)-CBC activated TRPA1; at 30 µM (-)-CBC produced an activation 50% of that of the reference agonist cinnamaldehyde (300 µM), 30 µM (+)-CBC activated TRPA1 to 38% of the cinnamaldehyde maximum. Discussion: It is unclear whether (-)-CBC is the sole or even the predominant enantiomer of CBC enzymatically synthesized in cannabis. This study shows that (-)-CBC is the active isomer at CB2 receptors, while both isomers activate TRPA1. The results suggest that medicinal preparations of CBC that target cannabinoid receptors would be most effective when (-)-CBC is the dominant isomer.

Cannabinoid receptor-2 agonist AM1241 attenuates myocardial ischemia-reperfusion-induced oxidative stress in rats via Nrf2/HO-1 pathway.

The cannabinoid receptor-2 agonist AM1241 exhibits notable cardioprotective effects against myocardial infarction, positioning it as a promising therapeutic candidate for cardiovascular disease. This study explores AM1241's protective role in myocardial ischemia-reperfusion (IR) injury and its association with the Nrf2/HO-1 pathway. In an established SD rat IR model, AM1241's impact on cardiac injury was assessed through echocardiography, 2,3,5-triphenyl tetrazolium chloride staining, and histological analysis. H9c2 cells underwent hypoxia-reoxygenation, with AM1241's influence on cell viability determined by the CCK-8 assay. Reactive oxygen species (ROS) production was measured using the DCFH-DA assay, and Nrf2 and HO-1 protein expressions were evaluated through immunofluorescence and Western blot. Myocardial ischemia-reperfusion injury (MIRI) increased infarct size, inflammatory cell presence, oxidative and nitrosative stress, impaired cardiac function, and elevated apoptosis rates. AM1241 mitigated these effects, enhancing cell viability, reducing ROS production, and upregulating Nrf2 and HO-1 expression. The antioxidant effect of AM1241 was inhibited by ML385 intervention. AM1241 attenuates oxidative stress, alleviates MIRI, and activates the Nrf2/HO-1 signaling pathway, underscoring its potential as a therapeutic strategy for myocardial ischemia-reperfusion injury.

Clinical and cognitive outcomes in first-episode psychosis: focus on the interplay between cannabis use and genetic variability in endocannabinoid receptors.

Research data show the impact of the endocannabinoid system on psychosis through its neurotransmission homeostatic functions. However, the effect of the endocannabinoid system genetic variability on the relationship between cannabis use and psychosis has been unexplored, even less in first-episode patients. Here, through a case-only design, we investigated the effect of cannabis use and the genetic variability of endocannabinoid receptors on clinical and cognitive outcomes in first-episode psychosis (FEP) patients. The sample comprised 50 FEP patients of European ancestry (mean age (sd) = 26.14 (6.55) years, 76% males), classified as cannabis users (58%) or cannabis non-users. Two Single Nucleotide Polymorphisms (SNP) were genotyped at the cannabinoid receptor type 1 gene (CNR1 rs1049353) and cannabinoid receptor type 2 gene (CNR2 rs2501431). Clinical (PANSS, GAF) and neuropsychological (WAIS, WMS, BADS) assessments were conducted. By means of linear regression models, we tested the main effect of cannabis use and its interaction with the polymorphic variants on the clinical and cognitive outcomes. First, as regards cannabis effects, our data showed a trend towards more severe positive symptoms (PANSS, p = 0.05) and better performance in manipulative abilities (matrix test-WAIS, p = 0.041) among cannabis users compared to non-users. Second, concerning the genotypic effects, the T allele carriers of the CNR1 rs1049353 presented higher PANSS disorganization scores than CC homozygotes (p = 0.014). Third, we detected that the observed association between cannabis and manipulative abilities is modified by the CNR2 polymorphism (p = 0.022): cannabis users carrying the G allele displayed better manipulative abilities than AA genotype carriers, while the cannabis non-users presented the opposite genotype-performance pattern. Such gene-environment interaction significantly improved the overall fit of the cannabis-only model (Δ-R2 = 8.4%, p = 0.019). Despite the preliminary nature of the sample, our findings point towards the role of genetic variants at CNR1 and CNR2 genes in the severity of the disorganized symptoms of first-episode psychosis and modulating cognitive performance conditional to cannabis use. This highlights the need for further characterization of the combined role of endocannabinoid system genetic variability and cannabis use in the understanding of the pathophysiology of psychosis.

Preclinical Assessment of a Cannabinoid CB2 Receptor Antagonist in a Murine Model of Cerebral Malaria

Malaria is a most important parasitic disease due to its highest impact worldwide. It results in around 200 million clinical cases and 0,5-1 million deaths per year, mainly due to cerebral malaria (CM), a life-threatening neurological syndrome that predominantly affects predominantly children under five years old. CM follows neurological alterations leading to the death if left untreated, and, even when it is treated, it is fatal in 15-20% of cases. Moreover, among the survivors, more than 10% of the children develop neurological sequelae. Consequently, there is an urgent need to find therapies to attenuate these neurological signs. Recent evidence has proposed the endocannabinoid system, which plays an important neuromodulatory function in the central nervous system (CNS), also including immunomodulation preferentially exerted by CB2 receptor. Previous studies have shown that the genetic ablation of this receptor improved mice survival against CM, suggesting a potential for the pharmacological treatment of CM with selective antagonists of this receptor. Considering this background, we investigated CM therapy by a classic CB2 antagonist SR144528 in a murine model of the disease. First, we carried out binding studies with SR144528 to confirm its pharmacodynamic profile (binding affinity [Ki] value = 2.34 ± 0.61 nM; and efficacy [IC50] = 96.17 ± 1.41 nM, at the CB2 receptor). Second, P. berghei ANKA infected C57BL/6 mice were treated daily with SR144528 and assessed for parasitemia growth and neurological alterations. 30% of the treated mice showed partial recovery of CM symptoms with 20% increased survival, but finally succumbing to hyperparasitemia and severe anemia. These preliminary preclinical results suggest that, although part of the CM course might be modulated by the pharmacological blockade of the CB2 receptor, other elements trigger the lethal outcome. Thus, while our hypothesis could not be completely validated in this CM model, we detail here all obtained results for further research.

A lactate-dependent shift of glycolysis mediates synaptic and cognitive processes in male mice

Astrocytes control brain activity via both metabolic processes and gliotransmission, but the physiological links between these functions are scantly known. Here we show that endogenous activation of astrocyte type-1 cannabinoid (CB1) receptors determines a shift of glycolysis towards the lactate-dependent production of d-serine, thereby gating synaptic and cognitive functions in male mice. Mutant mice lacking the CB1 receptor gene in astrocytes (GFAP-CB1-KO) are impaired in novel object recognition (NOR) memory. This phenotype is rescued by the gliotransmitter d-serine, by its precursor l-serine, and also by lactate and 3,5-DHBA, an agonist of the lactate receptor HCAR1. Such lactate-dependent effect is abolished when the astrocyte-specific phosphorylated-pathway (PP), which diverts glycolysis towards l-serine synthesis, is blocked. Consistently, lactate and 3,5-DHBA promoted the co-agonist binding site occupancy of CA1 post-synaptic NMDA receptors in hippocampal slices in a PP-dependent manner. Thus, a tight cross-talk between astrocytic energy metabolism and gliotransmission determines synaptic and cognitive processes.

Synthesis and Antitumour Evaluation of Tricyclic Indole-2-Carboxamides against Paediatric Brain Cancer Cells.

Antitumour properties of some cannabinoids (CB) have been reported in the literature as early as 1970s, however there is no clear consensus to date on the exact mechanisms leading to cancer cell death. The indole-based WIN 55,212-2 and SDB-001 are both known as potent agonists at both CB1 and CB2 receptors, yet we demonstrate herein that only the former can exert in vitro antitumour effects when tested against a paediatric brain cancer cell line KNS42. In this report, we describe the synthesis of novel 3,4-fused tricyclic indoles and evaluate their functional potencies at both cannabinoid receptors, as well as their abilities to inhibit the growth or proliferation of KNS42 cells. Compared to our previously reported indole-2-carboxamides, these 3,4-fused tricyclic indoles had either completely lost activities, or, showed moderate-to-weak antagonism at both CB1 and CB2 receptors. Compound 23 displayed the most potent antitumour properties among the series. Our results further support the involvement of non-CB pathways for the observed antitumour activities of amidoalkylindole-based cannabinoids, in line with our previous findings. Transcriptomic analysis comparing cells treated or non-treated with compound 23 suggested the observed antitumour effects of 23 are likely to result mainly from disruption of the FOXM1-regulated cell cycle pathways.