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Introduction: The contribution of Cannabinoid type 1 receptor (CB1) in mitochondrial energy transduction mechanisms and mitochondrial activities awaits deeper investigations. Our study aims to assess the impact of CB1 absence on the mitochondrial compartment in the liver, focusing on both functional aspects and remodeling processes. Methods: We used CB1−/− and CB1+/+ male mice. Cytochrome C Oxidase activity was determined polarographically. The expression and the activities of separated mitochondrial complexes and supercomplexes were performed by using Blue-Native Page, Western blotting and histochemical staining for in-gel activity. Key players of Mitochondrial Quality Control processes were measured using RT-qPCR and Western blotting. Liver fine sub-cellular ultrastructural features were analyzed by TEM analysis. Results and discussion: In the absence of CB1, several changes in the liver occur, including increased oxidative capacity, reduced complex I activity, enhanced complex IV activity, general upregulation of respiratory supercomplexes, as well as higher levels of oxidative stress. The mitochondria and cellular metabolism may be affected by these changes, increasing the risk of ROS-related damage. CB1−/− mice show upregulation of mitochondrial fusion, fission and biogenesis processes which suggests a dynamic response to the absence of CB1. Furthermore, oxidative stress disturbs mitochondrial proteostasis, initiating the mitochondrial unfolded protein response (UPRmt). We noted heightened levels of pivotal enzymes responsible for maintaining mitochondrial integrity, along with heightened expression of molecular chaperones and transcription factors associated with cellular stress reactions. Additionally, our discoveries demonstrate a synchronized reaction to cellular stress, involving both UPRmt and UPRER pathways. [ABSTRACT FROM AUTHOR]

Fibromyalgia is a chronic illness usually accompanied by long-lasting, general pain throughout the body, often accompanied by anxiety, depression, fatigue, and sleep disruption. Meanwhile, doctors and scientists have not entirely discovered detailed mechanisms; patients always have an exaggerated sensation to pervasive pain without satisfied medical service. Given the lack of knowledge on its underlying mechanism, current treatments aim to provide pain and/or symptom relief. The present study aimed to clarify the role of cannabinoid receptor 1 (CB1) signaling in a mouse fibromyalgia pain model. To develop the mouse fibromyalgia model, mice were subjected to intermittent cold stress (ICS). Our results indicated that mechanical (2.09 ± 0.09 g) and thermal hyperalgesia (4.77 ± 0.29 s), which were evaluated by von Frey and Hargraves' tests, were induced by ICS, suggesting successful modeling. The hurting replies were then provoked by electroacupuncture (EA) but not for sham EA mice. Further, in a Western blot analysis, we found significantly decreased CB1 protein levels in the thalamus, somatosensory cortex, and anterior cingulate cortex. In addition, the levels of pain-related protein kinases and transcription factor were increased. Treatment with EA reliably increased CB1 expression in various brain regions sequentially alleviated by nociceptive mediators. Furthermore, the administration of a CB1 agonist significantly attenuated fibromyalgia pain, reversed EA analgesia by the CB1 antagonist, and further reversed the chemogenetic inhibition of SSC. Our innovative findings evidence the role of CB1 signaling in the interaction of EA and fibromyalgia, suggesting its potential for clinical trials and as a treatment target. [ABSTRACT FROM AUTHOR]

Background and Purpose: Transient hypofunction of the NMDA receptor represents a convergence point for the onset and further development of psychiatric disorders, including schizophrenia. Although the cumulative evidence indicates dysregulation of the hippocampal formation in schizophrenia, the integrity of the synaptic transmission and plasticity conveyed by the somatosensorial inputs to the dentate gyrus, the perforant pathway synapses, have barely been explored in this pathological condition. Experimental Approach: We identified a series of synaptic alterations of the lateral and medial perforant paths in animals postnatally treated with the NMDA antagonist MK‐801. This dysregulation suggests decreased cognitive performance, for which the dentate gyrus is critical. Key Results: We identified alterations in the synaptic properties of the lateral and medial perforant paths to the dentate gyrus synapses in slices from MK‐801‐treated animals. Altered glutamate release and decreased synaptic strength precede an impairment in the induction and expression of long‐term potentiation (LTP) and CB1 receptor‐mediated long‐term depression (LTD). Remarkably, by inhibiting the degradation of 2‐arachidonoylglycerol (2‐AG), an endogenous ligand of the CB1 receptor, we restored the LTD in animals treated with MK‐801. Additionally, we showed for the first time, that spatial discrimination, a cognitive task that requires dentate gyrus integrity, is impaired in animals exposed to transient hypofunction of NMDA receptors. Conclusion and Implications: Dysregulation of glutamatergic transmission and synaptic plasticity from the entorhinal cortex to the dentate gyrus has been demonstrated, which may explain the cellular dysregulations underlying the altered cognitive processing in the dentate gyrus associated with schizophrenia. [ABSTRACT FROM AUTHOR]

The endocannabinoid system interacts with the reward system to modulate responsiveness to natural reinforcers, as well as drugs of abuse. Previous preclinical studies suggested that direct blockade of CB1 cannabinoid receptors (CB1R) could be leveraged as a potential pharmacological approach to treat substance use disorder, but this strategy failed during clinical trials due to severe psychiatric side effects. Alternative strategies have emerged to circumvent the side effects of direct CB1 binding through the development of allosteric modulators. We hypothesized that negative allosteric modulation of CB1R signalling would reduce the reinforcing properties of morphine and decrease behaviours associated with opioid misuse. By employing intravenous self‐administration in mice, we studied the effects of GAT358, a functionally‐biased CB1R negative allosteric modulator (NAM), on morphine intake, relapse‐like behaviour and motivation to work for morphine infusions. GAT358 reduced morphine infusion intake during the maintenance phase of morphine self‐administration under a fixed ratio 1 schedule of reinforcement. GAT358 also decreased morphine‐seeking behaviour after forced abstinence. Moreover, GAT358 dose dependently decreased the motivation to obtain morphine infusions under a progressive ratio schedule of reinforcement. Strikingly, GAT358 did not affect the motivation to work for food rewards in an identical progressive ratio task, suggesting that the effect of GAT358 in decreasing opioid self‐administration was reward specific. Furthermore, GAT58 did not produce motor ataxia in the rotarod test. Our results suggest that CB1R NAMs reduced the reinforcing properties of morphine and could represent a viable therapeutic route to safely decrease misuse of opioids. [ABSTRACT FROM AUTHOR]

Datura stramonium, Moringa oleifera and Carica papaya are common plants in Nigeria that have been reported to possess some psychoactive effects. However, the interactions of their alkaloids with the molecular targets of common psychoactive compounds are not well established. This study assessed the interactive potentials of alkaloids from these plants with a4ß2 nicotinic acetylcholine receptor (a4ß2 nAChR) of nicotine from Nicotiana tabacum and the cannabinoid receptor 1 (CB1) of delta-9-tetrahydrocannabinol (THC) from Cannabis sativa. Protein structures were retrieved from Protein Data Bank while PubChem was used to obtain ligand structures. Molecular docking using UCSF Chimera determined the binding affinity of protein-ligand complexes, followed by molecular dynamics simulations to evaluate root mean square deviation and radius of gyration. ADMET analysis was performed using SwissADME and ProTox-II. Notably, apoatropine, hyoscyamine, and 3â,6â-ditigloyloxytropane from D. stramonium exhibited stronger a4ß2 nAChR binding effects, compared to nicotine, and had CB1 binding affinities similar to THC. Among these high-affinity binding compounds, apoatropine maintained the most stable and compact structural conformation, relative to nicotine and THC. ADMET analysis indicated propitious physicochemical and drug-like properties for all plantbased alkaloids except N, a-L-rhamnopyranosyl vincosamide from M. oleifera and apoatropine, which were predicted to be carcinogenic. Additionally, over 50% of the plant-based alkaloids assessed are blood-brain barrier permeant, implying their propensity to mediate CNS effects. It is pertinent to regulate the use of these plants, particularly in tropical regions like Nigeria, where they are widely cultivated, consumed, and likely explored for recreational purposes based on their psychoactive effects. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

IntroductionThe contribution of Cannabinoid type 1 receptor (CB1) in mitochondrial energy transduction mechanisms and mitochondrial activities awaits deeper investigations. Our study aims to assess the impact of CB1 absence on the mitochondrial compartment in the liver, focusing on both functional aspects and remodeling processes.MethodsWe used CB1−/− and CB1+/+ male mice. Cytochrome C Oxidase activity was determined polarographically. The expression and the activities of separated mitochondrial complexes and supercomplexes were performed by using Blue-Native Page, Western blotting and histochemical staining for in-gel activity. Key players of Mitochondrial Quality Control processes were measured using RT-qPCR and Western blotting. Liver fine sub-cellular ultrastructural features were analyzed by TEM analysis.Results and discussionIn the absence of CB1, several changes in the liver occur, including increased oxidative capacity, reduced complex I activity, enhanced complex IV activity, general upregulation of respiratory supercomplexes, as well as higher levels of oxidative stress. The mitochondria and cellular metabolism may be affected by these changes, increasing the risk of ROS-related damage. CB1−/− mice show upregulation of mitochondrial fusion, fission and biogenesis processes which suggests a dynamic response to the absence of CB1. Furthermore, oxidative stress disturbs mitochondrial proteostasis, initiating the mitochondrial unfolded protein response (UPRmt). We noted heightened levels of pivotal enzymes responsible for maintaining mitochondrial integrity, along with heightened expression of molecular chaperones and transcription factors associated with cellular stress reactions. Additionally, our discoveries demonstrate a synchronized reaction to cellular stress, involving both UPRmt and UPRER pathways.

Fibromyalgia is a chronic illness usually accompanied by long-lasting, general pain throughout the body, often accompanied by anxiety, depression, fatigue, and sleep disruption. Meanwhile, doctors and scientists have not entirely discovered detailed mechanisms; patients always have an exaggerated sensation to pervasive pain without satisfied medical service. Given the lack of knowledge on its underlying mechanism, current treatments aim to provide pain and/or symptom relief. The present study aimed to clarify the role of cannabinoid receptor 1 (CB1) signaling in a mouse fibromyalgia pain model. To develop the mouse fibromyalgia model, mice were subjected to intermittent cold stress (ICS). Our results indicated that mechanical (2.09 ± 0.09 g) and thermal hyperalgesia (4.77 ± 0.29 s), which were evaluated by von Frey and Hargraves’ tests, were induced by ICS, suggesting successful modeling. The hurting replies were then provoked by electroacupuncture (EA) but not for sham EA mice. Further, in a Western blot analysis, we found significantly decreased CB1 protein levels in the thalamus, somatosensory cortex, and anterior cingulate cortex. In addition, the levels of pain-related protein kinases and transcription factor were increased. Treatment with EA reliably increased CB1 expression in various brain regions sequentially alleviated by nociceptive mediators. Furthermore, the administration of a CB1 agonist significantly attenuated fibromyalgia pain, reversed EA analgesia by the CB1 antagonist, and further reversed the chemogenetic inhibition of SSC. Our innovative findings evidence the role of CB1 signaling in the interaction of EA and fibromyalgia, suggesting its potential for clinical trials and as a treatment target.

One of the hallmarks of Parkinson's disease (PD) is the alteration in the expression and function of NMDA receptor (NMDAR) and cannabinoid receptor 1 (CB1R). The presence of CB1R-NMDAR complexes has been described in neuronal primary cultures. The activation of CB1R in CB1R-NMDAR complexes was suggested to counteract the detrimental NMDAR overactivation in an AD mice model. Thus, we aimed to explore the role of this receptor complex in PD. By using Bioluminescence Resonance Energy Transfer (BRET) assay, it was demonstrated that α-synuclein induces a reorganization of the CB1R-NMDAR complex in transfected HEK-293T cells. Moreover, α-synuclein treatment induced a decrease in the cAMP and MAP kinase (MAPK) signaling of both CB1R and NMDAR not only in transfected cells but also in neuronal primary cultures. Finally, the interaction between CB1R and NMDAR was studied by Proximity Ligation Assay (PLA) in neuronal primary cultures, where it was observed that the expression of CB1R-NMDAR complexes was decreased upon α-synuclein treatment. These results point to a role of CB1R-NMDAR complexes as a new therapeutic target in Parkinson's disease. [ABSTRACT FROM AUTHOR]

In childhood, retinoblastoma (RB) is the most common primary tumor in the eye. Long term therapeutic management with etoposide of this life-threatening condition may have diminishing effectiveness since RB cells can develop cytostatic resistance to this drug. To determine whether changes in receptor-mediated control of Ca2+ signaling are associated with resistance development, fluorescence calcium imaging, semi-quantitative RT-qPCR analyses, and trypan blue dye exclusion staining patterns are compared in WERI-ETOR (etoposide-insensitive) and WERI-Rb1 (etoposide-sensitive) cells. The cannabinoid receptor agonist 1 (CNR1) WIN55,212-2 (40 µM), or the transient receptor potential melastatin 8 (TRPM8) agonist icilin (40 µM) elicit similar large Ca2+ transients in both cell line types. On the other hand, NGF (100 ng/mL) induces larger rises in WERI-ETOR cells than in WERI-Rb1 cells, and its lethality is larger in WERI-Rb1 cells than in WERI-ETOR cells. NGF and WIN55,212-2 induced additive Ca2+ transients in both cell types. However, following pretreatment with both NGF and WIN55,212-2, TRPM8 gene expression declines and icilin-induced Ca2+ transients are completely blocked only in WERI-ETOR cells. Furthermore, CNR1 gene expression levels are larger in WERI-ETOR cells than those in WERI-Rb1 cells. Therefore, the development of etoposide insensitivity may be associated with rises in CNR1 gene expression, which in turn suppress TRPM8 gene expression through crosstalk. [ABSTRACT FROM AUTHOR]

AIM: To explore the modulation of 2-arachidonoylglycerol (2-AG) on A-type potassium channels injured by kainic acid (KA) and its molecular mechanism. METHODS: Primary cultured caudate nucleus (CN) neurons were treated with KA to establish a neuroexcitatory toxicity model. Whole-cell patch clamp recording was performed to observe the changes of electrical activity of A-type potassium channels induced by KA-induced excitatory toxicity and 2-AG-mediated neuroprotective effect. RESULTS: In cultured CN neurons, patch clamp experiments confirmed that KA significantly decreased the A-type potassium channel current (Iʌ) density and changed the electrical function of CN neurons: the slope (k) of inactivation curve and the recovery time constant (τ) after inactivation of A-type potassium channels in CN neurons were significantly increased. The experiments showed that the increase in 2-AG level, whether using 2-AG directly or application of monoacylglycerol lipase inhibitor URB602 to decrease 2-AG metabolism and increase 2-AG level indirectly, inhibited the KA-induced reduction of Iʌ density and the changes of electrical activity of A-type potassium channels through cannabinoid receptor 1 (CB1R) : 2-AG effectively antagonized the KA-induced increases in τ value and k value for inactivation of A-type potassium channels, which accelerated the recovery process after inactivation of the channels. CONCLUSION: The changes of the electrical characteristics of A-type potassium channels may be one of the mechanisms of KA-induced excitotoxic injury of CN neurons. The 2-AG plays a neuroprotective role in KA-induced neuroexcitatory toxic model by regulating the function of A-type potassium channels through CB1R. [ABSTRACT FROM AUTHOR]

The cannabinoid receptor 1 (CB1R) plays a pivotal role in regulating various physiopathological processes, thus positioning itself as a promising and sought-after therapeutic target. However, the search for specific and effective CB1R ligands has been challenging, prompting the exploration of drug repurposing (DR) strategies. In this study, we present an innovative DR approach that combines computational screening and experimental validation to identify potential Food and Drug Administration (FDA)-approved compounds that can interact with the CB1R. Initially, a large-scale virtual screening was conducted using molecular docking simulations, where a library of FDA-approved drugs was screened against the CB1R's three-dimensional structures. This in silico analysis allowed us to prioritize compounds based on their binding affinity through two different filters. Subsequently, the shortlisted compounds were subjected to in vitro assays using cellular and biochemical models to validate their interaction with the CB1R and determine their functional impact. Our results reveal FDA-approved compounds that exhibit promising interactions with the CB1R. These findings open up exciting opportunities for DR in various disorders where CB1R signaling is implicated. In conclusion, our integrated computational and experimental approach demonstrates the feasibility of DR for discovering CB1R modulators from existing FDA-approved compounds. By leveraging the wealth of existing pharmacological data, this strategy accelerates the identification of potential therapeutics while reducing development costs and timelines. The findings from this study hold the potential to advance novel treatments for a range of CB1R -associated diseases, presenting a significant step forward in drug discovery research. [ABSTRACT FROM AUTHOR]

Cerebral metabolic abnormalities are common in neurodegenerative diseases. Previous studies have shown that mitochondrial damage alters ATP production and increases reactive oxygen species (ROS) release which may contribute to neurodegeneration. In the present study, we investigated the neuroprotective effects of cannabidiol (CBD), a non-psychoactive component derived from marijuana (Cannabis sativa L.), on astrocytic bioenergetic balance in a primary cell culture model of lipopolysaccharide (LPS)-induced neurotoxicity. Astrocytic metabolic profiling using an extracellular flux analyzer demonstrated that CBD decreases mitochondrial proton leak, increased spare respiratory capacity and coupling efficiency in LPS-stimulated astrocytes. Simultaneously, CBD increased astrocytic glycolytic capacity and glycolysis reserve in a cannabinoid receptor type 1 (CB1)-dependent manner. CBD-restored metabolic changes were correlated with a significant decrease in the pro-inflammatory cytokines tumor necrosis factor α (TNFα) and interleukin-6 (IL-6) concentration and reduction of ROS production in LPS-stimulated astrocytes. These results suggest that CBD may inhibit LPS-induced metabolic impairments and inflammation by enhancing astrocytic metabolic glycolysis versus oxidative phosphorylation through its action on CB1 receptors. The present findings suggest CBD as a potential anti-inflammatory treatment in metabolic pathologies and highlight a possible role for the cannabinoidergic system in the modulation of mitochondrial oxidative stress. CBD enhances mitochondrial bioenergetic profile, attenuates proinflammatory cytokines release, and ROS overproduction of astrocytes stimulated by LPS. These effects are not mediated directly by CB1 receptors, while these receptors seem to have a key role in the anti-inflammatory response of the endocannabinoid system on astrocytes, as their specific inhibition by SR141716A led to increased pro-inflammatory cytokines release and ROS production. The graphical abstract is created with BioRender.com. [ABSTRACT FROM AUTHOR]

In individuals with type 2 diabetes mellitus (T2DM), the cannabinoid receptor 1 (CNR1) gene polymorphism has been linked to diabetic nephropathy (DN). Different renal disorders, including DN, have been found to alter cannabinoid (CB) receptor expression and activation. This cross-sectional study aimed to investigate the relationship between CNR1 rs1776966256 and rs1243008337 genetic variants and the risk of developing DN in Iraqi patients with T2DM. The study included 100 patients with T2DM, divided into two groups: 50 with DN and 50 without DN. Genotyping of CNR1 rs1776966256 and rs1243008337 polymorphisms was conducted using PCR in DN patients and control samples. The distribution of rs1776966256 and rs1243008337 genotypes and alleles between the two groups revealed statistically significant differences. The frequencies of the GG and AG genotypes of CNR1 rs1776966256 were significantly different between DN patients and the control group. Additionally, compared to the A allele, the G allele of this polymorphism was linked to a higher incidence of DN (p=0.0001). Patients with the genetic polymorphism rs1243008337 had higher genotypes of CC and AC and were more likely to develop DN in the polymorphism genotype than the wild genotype. Additionally, compared to the A allele, the C allele was linked to a higher chance of developing DN (p=0.0001). Both rs1776966256 and rs1243008337 polymorphisms were correlated with the development of diabetic nephropathy. [ABSTRACT FROM AUTHOR]

Neuroinflammation is a complex biological process that typically originates as a protective response in the brain. This inflammatory process is triggered by the release of pro-inflammatory substances like cytokines, prostaglandins, and reactive oxygen and nitrogen species from stimulated endothelial and glial cells, including those with pro-inflammatory functions, in the outer regions. While neuronal inflammation is common in various central nervous system disorders, the specific inflammatory pathways linked with different immune-mediated cell types and the various factors influencing the blood-brain barrier significantly contribute to disease-specific characteristics. The endocannabinoid system consists of cannabinoid receptors, endogenous cannabinoids, and enzymes responsible for synthesizing and metabolizing endocannabinoids. The primary cannabinoid receptor is CB1, predominantly found in specific brain regions such as the brainstem, cerebellum, hippocampus, and cortex. The presence of CB2 receptors in certain brain components, like cultured cerebellar granular cells, Purkinje fibers, and microglia, as well as in the areas like the cerebral cortex, hippocampus, and cerebellum is also evidenced by immunoblotting assays, radioligand binding, and autoradiography studies. Both CB1 and CB2 cannabinoid receptors exhibit noteworthy physiological responses and possess diverse neuromodulatory capabilities. This review primarily aims to outline the distribution of CB1 and CB2 receptors across different brain regions and explore their potential roles in regulating neuroinflammatory processes. [ABSTRACT FROM AUTHOR]

The rostral agranular insular cortex (RAIC) has been associated with pain modulation. Although the endogenous cannabinoid system (eCB) has been shown to regulate chronic pain, the roles of eCBs in the RAIC remain elusive under the neuropathic pain state. Neuropathic pain was induced in C57BL/6 mice by common peroneal nerve (CPN) ligation. The roles of the eCB were tested in the RAIC of ligated CPN C57BL/6J mice, glutamatergic, or GABAergic neuron cannabinoid receptor 1 (CB1R) knockdown mice with the whole-cell patch-clamp and pain behavioral methods. The E/I ratio (amplitude ratio between mEPSCs and mIPSCs) was significantly increased in layer V pyramidal neurons of the RAIC in CPN-ligated mice. Depolarization-induced suppression of inhibition but not depolarization-induced suppression of excitation in RAIC layer V pyramidal neurons were significantly increased in CPN-ligated mice. The analgesic effect of ACEA (a CB1R agonist) was alleviated along with bilateral dorsolateral funiculus lesions, with the administration of AM251 (a CB1R antagonist), and in CB1R knockdown mice in GABAergic neurons, but not glutamatergic neurons of the RAIC. Our results suggest that CB1R activation reinforces the function of the descending pain inhibitory pathway via reducing the inhibition of glutamatergic layer V neurons by GABAergic neurons in the RAIC to induce an analgesic effect in neuropathic pain. [ABSTRACT FROM AUTHOR]

In the central nervous system (CNS), cannabinoid receptor 1 (CB1R) is preferentially expressed in axons where it has a unique property, namely resistance to agonist-driven endocytosis. This review aims to summarize what we know about molecular mechanisms of CB1R cell surface stability in axonal compartments, how these impact CB1R signaling, and to consider their physiological consequences. This review then focuses on a potential candidate for maintaining axonal CB1R at the cell surface, Src homology 3-domain growth factor receptor-bound 2-like endophilin interacting protein 1 (SGIP1). SGIP1 may contribute to the polarized distribution of CB1R and modify its signaling in axons. In addition, deletion of SGIP1 results in discrete behavioral changes in modalities controlled by the endocannabinoid system in vivo. Several drugs acting directly via CB1R have important therapeutic potential, however their adverse effects limit their clinical use. Future studies might reveal chemical approaches to target the SGIP1-CB1R interaction, with the aim to exploit the endocannabinoid system pharmaceutically in a discrete way, with minimized undesired consequences. [ABSTRACT FROM AUTHOR]

Drugs of abuse induce cell type‐specific adaptations in D1‐ and D2‐medium spiny neurons (MSNs) in the nucleus accumbens core (NAcore) that can bias signalling towards D1‐MSNs and enhance relapse vulnerability. Whether Δ9‐tetrahydrocannabinol (THC) use initiates similar neuroadaptations is unknown. D1‐ and D2‐Cre transgenic rats were transfected with Cre‐dependent reporters and trained to self‐administer THC + cannabidiol (THC + CBD). After extinction training spine morphology, glutamate transmission, CB1R function and cFOS expression were quantified. We found that extinction from THC + CBD induced a loss of large spine heads in D1‐ but not D2‐MSNs and commensurate reductions in glutamate synaptic transmission. Also, presynaptic CB1R function was impaired selectively at glutamatergic synapses on D1‐MSNs, which augmented the capacity to potentiate glutamate transmission. Using cFOS expression as an activity marker, we found no change after extinction but increased cFOS expression in D1‐MSNs after cue‐induced drug seeking. Contrasting D1‐MSNs, CB1R function and glutamate synaptic transmission on D2‐MSN synapses were unaffected by THC + CBD use. However, cFOS expression was decreased in D2‐MSNs of THC + CBD‐extinguished rats and was restored after drug seeking. Thus, CB1R adaptations in D1‐MSNs partially predicted neuronal activity changes, posing pathway specific modulation of eCB signalling in D1‐MSNs as a potential treatment avenue for cannabis use disorder (CUD). [ABSTRACT FROM AUTHOR]

Background: Electroacupuncture (EA) can effectively relieve visceral hypersensitivity (VH). However, its mechanisms are still unclear. Objective: To investigate the impact of EA on VH caused by ileitis, and whether EA relieves VH by modulating the endogenous cannabinoid system (ECS). Methods: Thirty male native goats were randomly divided into a saline-treated control group (Saline, n = 9) and three 2,4,6-trinitro-benzenesulfonic acid (TNBS)-treated VH model groups that underwent injection of TNBS into the ileal wall to induce VH and remained untreated (TNBS, n = 9) or received six sessions of EA (for 30 min every 3 days) (TNBS + EA, n = 6) or sham acupuncture (TNBS + Sham, n = 6). The visceromotor response (VMR) to colorectal distention (CRD) was measured after each EA treatment. Three goats in the Saline/TNBS groups were euthanized after 7 days for histopathological examination; the remaining 24 (n = 6/group) underwent sampling of the ileal wall, T11 spinal cord and brain nuclei/areas related to visceral regulation and ascending pain modulation system on day 22. Expression of cannabinoid receptor 1 (CB1R), fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL) was detected by immunohistochemistry. Results: VMR to CRD was greater in TNBS-treated goats than in saline-treated goats (p < 0.01) from day 7 to 22. After day 7, EA-treated goats showed a decreased (p < 0.05) VMR compared with untreated TNBS-exposed goats. TNBS treatment decreased CB1R and increased FAAH and MAGL expression in the ileum and related nuclei/areas; this was reversed by EA. Conclusion: EA ameliorates VH, probably by regulating the ECS in the intestine and nuclei/areas related to visceral regulation and descending pain modulation systems. [ABSTRACT FROM AUTHOR]

Background: Diabetic Kidney Disease (DKD) is a significant challenge in healthcare. However, there are currently no reliable biomarkers for renal impairment diagnosis, prognosis, or staging in DKD patients. CircRNAs and microRNAs have emerged as noninvasive and efficient biomarkers. Methods: We explored Cannabinoid receptor 1 (CNR1), C reactive protein (CRP), hsa_circ_ 0000146 and 0000072, and hsa-miR-21 and 495 as diagnostic biomarkers in DKD. The serum concentrations of CRP and CNR1 were measured using ELISA. Rt-qPCR was used to evaluate the expression levels of CNR1, circRNAs, and miRNAs in 55 controls, 55 type 2 diabetes mellitus patients, and 55 DKD patients. Their diagnostic value was determined by their ROC curve. KEGG pathway was used to predict the functional mechanism of the circRNA's target genes. Results: DKD patients exhibited a significant increase in CRP and CNR1 levels and the expression of miR-21 and 495. The expression levels of circ_0000146 and 0000072 decreased in DKD patients. ROC analysis revealed that circRNAs and miRNAs alone or CNR1 and CRP have significant diagnostic potential. The functional prediction results showed the involvement of hsa_circ_0000146 and 0000072 in various pathways that regulate DKD. Conclusions: Therefore, the examined circRNAs and miRNAs may represent a novel noninvasive biomarker for diagnosing and staging DKD.

Emerging evidence indicates that the endogenous cannabinoid system modulates the behavioral and physiological effects of nicotine. Fatty acid-binding proteins (FABPs) are among the primary intracellular trafficking mechanisms of endogenous cannabinoids, such as anandamide. To this end, changes in FABP expression may similarly impact the behavioral manifestations associated with nicotine, particularly its addictive properties. FABP5+/+ and FABP5−/− mice were tested for nicotine-conditioned place preference (CPP) at two different doses (0.1 or 0.5 mg/kg). The nicotine-paired chamber was assigned as their least preferred chamber during preconditioning. Following 8 days of conditioning, the mice were injected with either nicotine or saline. The mice were allowed to access to all the chambers on the test day, and their times spent in the drug chamber on the preconditioning versus the test days were used to examine the drug preference score. The CPP results showed that the FABP5−/− mice displayed a higher place preference for 0.1 mg/kg nicotine than the FABP5+/+ mice, while no CPP difference was observed for 0.5 mg/kg nicotine between the genotypes. In conclusion, FABP5 plays an important role in regulating nicotine place preference. Further research is warranted to identify the precise mechanisms. The results suggest that dysregulated cannabinoid signaling may impact nicotine-seeking behavior.

Objective: The aim of this study was to identify and validate the reference genes in cultured human odontoblasts to quantify their cannabinoid receptor transcripts. Methods: The most stably transcribed genes in cultured human odontoblast cells were identified using the RefGenes tool and were selected for real-time polymerase chain reaction (PCR) amplification. Human odontoblast cells were differentiated from mesenchymal stem cells using a transforming growth factor-β-supplemented differentiation medium, and total RNA was purified. Reverse transcription-quantitative PCR and relative quantification analyses were performed using the Schefe's method. The relative expression dataset was analyzed to select the most stable genes. Results: The analysis showed that the transcripts of cholinergic receptor nicotinic beta 2 subunit, LIM homeobox transcription factor 1 beta, and family with sequence similarity 223 member B presented the lowest standard deviation (SD) in expression (SD: 0.2, 0.17, and 0.16, respectively). These genes showed similar expression levels as the target genes (cannabinoid receptors). Significant differences were found in the relative expression levels of cannabinoid receptors using the selected genes compared to those calculated using beta actin transcripts as references (p

Endocannabinoid signaling plays crucial roles in human physiology in the function of multiple systems. The two cannabinoid receptors, CB1 and CB2, are cell membrane proteins that interact with both exogenous and endogenous bioactive lipid ligands, or endocannabinoids. Recent evidence has established that endocannabinoid signaling operates within the human kidney, as well as suggests the important role it plays in multiple renal pathologies. CB1, specifically, has been identified as the more prominent ECS receptor within the kidney, allowing us to place emphasis on this receptor. The activity of CB1 has been repeatedly shown to contribute to both diabetic and non-diabetic chronic kidney disease (CKD). Interestingly, recent reports of acute kidney injury (AKI) have been attributed to synthetic cannabinoid use. Therefore, the exploration of the ECS, its receptors, and its ligands can help provide better insight into new methods of treatment for a range of renal diseases. This review explores the endocannabinoid system, with a focus on its impacts within the healthy and diseased kidney. [ABSTRACT FROM AUTHOR]

The endocannabinoid system (ECS) refers to a complex cell-signaling system highly conserved among species formed by numerous receptors, lipid mediators (endocannabinoids) and synthetic and degradative enzymes. It is widely distributed throughout the body including the CNS, where it participates in synaptic signaling, plasticity and neurodevelopment. Besides, the olfactory ensheathing glia (OEG) present in the olfactory system is also known to play an important role in the promotion of axonal growth and/or myelination. Therefore, both OEG and the ECS promote neurogenesis and oligodendrogenesis in the CNS. Here, we investigated if the ECS is expressed in cultured OEG, by assessing the main markers of the ECS through immunofluorescence, western blotting and qRT-PCR and quantifying the content of endocannabinoids in the conditioned medium of these cells. After that, we investigated whether the production and release of endocannabinoids regulate the differentiation of oligodendrocytes co-cultured with hippocampal neurons, through Sholl analysis in oligodendrocytes expressing O4 and MBP markers. Additionally, we evaluated through western blotting the modulation of downstream pathways such as PI3K/Akt/mTOR and ERK/MAPK, being known to be involved in the proliferation and differentiation of oligodendrocytes and activated by CB1, which is the major endocannabinoid responsive receptor in the brain. Our data show that OEG expresses key genes of the ECS, including the CB1 receptor, FAAH and MAGL. Besides, we were able to identify AEA, 2-AG and AEA related mediators palmitoylethanolamide (PEA) and oleoylethanolamide (OEA), in the conditioned medium of OEG cultures. These cultures were also treated with URB597 10-9 M, a FAAH selective inhibitor, or JZL184 10-9 M, a MAGL selective inhibitor, which led to the increase in the concentrations of OEA and 2-AG in the conditioned medium. Moreover, we found that the addition of OEG conditioned medium (OEGCM) enhanced the complexity of oligodendrocyte process branching in hippocampal mixed cell cultures and that this effect was inhibited by AM251 10-6 M, a CB1 receptor antagonist. However, treatment with the conditioned medium enriched with OEA or 2-AG did not alter the process branching complexity of premyelinating oligodendrocytes, while decreased the branching complexity in mature oligodendrocytes. We also observed no change in the phosphorylation of Akt and ERK 44/42 in any of the conditions used. In conclusion, our data show that the ECS modulates the number and maturation of oligodendrocytes in hippocampal mixed cell cultures. [ABSTRACT FROM AUTHOR]

In the central nervous system (CNS), cannabinoid receptor 1 (CB1R) is preferentially expressed in axons where it has a unique property, namely resistance to agonist-driven endocytosis. This review aims to summarize what we know about molecular mechanisms of CB1R cell surface stability in axonal compartments, how these impact CB1R signaling, and to consider their physiological consequences. This review then focuses on a potential candidate for maintaining axonal CB1R at the cell surface, Src homology 3-domain growth factor receptor-bound 2-like endophilin interacting protein 1 (SGIP1). SGIP1 may contribute to the polarized distribution of CB1R and modify its signaling in axons. In addition, deletion of SGIP1 results in discrete behavioral changes in modalities controlled by the endocannabinoid system in vivo. Several drugs acting directly via CB1R have important therapeutic potential, however their adverse effects limit their clinical use. Future studies might reveal chemical approaches to target the SGIP1-CB1R interaction, with the aim to exploit the endocannabinoid system pharmaceutically in a discrete way, with minimized undesired consequences.

Introduction: This study aimed to identify the effect of electroacupuncture (EA) treatment on post-stroke depression (PSD) and explore whether cannabinoid receptor 1 (CB1R)-mediated mitochondrial biogenesis accounts for the treatment effect of EA. Methods: The PSD mouse model was induced by a consecutive 14-day chronic unpredictable stress operation after 7 days of recovery from the bilateral common carotid artery occlusion surgery. Either EA treatment or sham stimulation was performed for 14 consecutive days from Day 7 after the BCCAO operation. Subjects' PSD-like behaviors were tested via open field test, sucrose preference test, novelty suppressed feeding test, tail suspension test, and forced swim test, and subjects' cognitive function was examined using Y-maze and novelty object recognition test. In addition, the levels of CB1R, mitochondrial biogenesis-related proteins (nuclear transcription factor 1, NRF1; mitochondrial transcription factor A, TFAM), proteins related to mitochondrial function (Cytochrome C, Cyto C; AIF, COX IV), and mitochondrial DNA were measured. To elucidate the role of CB1R in EA treatment, CB1R antagonists AM251 and CB1R-shRNA were given to mice before EA treatment. Likewise, subjects' depressive-like behaviors, cognitive function, mitochondrial function, and mitochondrial biogenesis were examined after the PSD procedure. Results: It has been showed that EA successfully ameliorated depressive-like behaviors, improved cognitive dysfunctions, and upregulated CB1R, NRF1 and TFAM expressions. However, the supplementation of AM251 and CB1R-shRNA blocked the antidepressant-like effects generated by EA, and EA failed to improve cognitive dysfunction, upregulate CB1R protein expression, and increase mitochondrial function and biogenesis. Conclusion: Altogether, these results indicated that EA ameliorated PSD-like behaviors in mice, improved cognitive dysfunctions after PSD, and promoted mitochondrial biogenesis by activating CB1R, a novel mechanism underlying EA's antidepressant-like effects in treating PSD. [ABSTRACT FROM AUTHOR]

Background: Diabetic Kidney Disease (DKD) is a significant challenge in healthcare. However, there are currently no reliable biomarkers for renal impairment diagnosis, prognosis, or staging in DKD patients. CircRNAs and microRNAs have emerged as noninvasive and efficient biomarkers. Methods: We explored Cannabinoid receptor 1 (CNR1), C reactive protein (CRP), hsa_circ_ 0000146 and 0000072, and hsa-miR-21 and 495 as diagnostic biomarkers in DKD. The serum concentrations of CRP and CNR1 were measured using ELISA. Rt-qPCR was used to evaluate the expression levels of CNR1, circRNAs, and miRNAs in 55 controls, 55 type 2 diabetes mellitus patients, and 55 DKD patients. Their diagnostic value was determined by their ROC curve. KEGG pathway was used to predict the functional mechanism of the circRNA's target genes. Results: DKD patients exhibited a significant increase in CRP and CNR1 levels and the expression of miR-21 and 495. The expression levels of circ_0000146 and 0000072 decreased in DKD patients. ROC analysis revealed that circRNAs and miRNAs alone or CNR1 and CRP have significant diagnostic potential. The functional prediction results showed the involvement of hsa_circ_0000146 and 0000072 in various pathways that regulate DKD. Conclusions: Therefore, the examined circRNAs and miRNAs may represent a novel noninvasive biomarker for diagnosing and staging DKD. [ABSTRACT FROM AUTHOR]

Emerging evidence indicates that the endogenous cannabinoid system modulates the behavioral and physiological effects of nicotine. Fatty acid-binding proteins (FABPs) are among the primary intracellular trafficking mechanisms of endogenous cannabinoids, such as anandamide. To this end, changes in FABP expression may similarly impact the behavioral manifestations associated with nicotine, particularly its addictive properties. FABP5+/+ and FABP5−/− mice were tested for nicotine-conditioned place preference (CPP) at two different doses (0.1 or 0.5 mg/kg). The nicotine-paired chamber was assigned as their least preferred chamber during preconditioning. Following 8 days of conditioning, the mice were injected with either nicotine or saline. The mice were allowed to access to all the chambers on the test day, and their times spent in the drug chamber on the preconditioning versus the test days were used to examine the drug preference score. The CPP results showed that the FABP5−/− mice displayed a higher place preference for 0.1 mg/kg nicotine than the FABP5+/+ mice, while no CPP difference was observed for 0.5 mg/kg nicotine between the genotypes. In conclusion, FABP5 plays an important role in regulating nicotine place preference. Further research is warranted to identify the precise mechanisms. The results suggest that dysregulated cannabinoid signaling may impact nicotine-seeking behavior. [ABSTRACT FROM AUTHOR]

The cannabinoid receptor 1 (CB1R) plays a pivotal role in regulating various physiopathological processes, thus positioning itself as a promising and sought-after therapeutic target. However, the search for specific and effective CB1R ligands has been challenging, prompting the exploration of drug repurposing (DR) strategies. In this study, we present an innovative DR approach that combines computational screening and experimental validation to identify potential Food and Drug Administration (FDA)-approved compounds that can interact with the CB1R. Initially, a large-scale virtual screening was conducted using molecular docking simulations, where a library of FDA-approved drugs was screened against the CB1R’s three-dimensional structures. This in silico analysis allowed us to prioritize compounds based on their binding affinity through two different filters. Subsequently, the shortlisted compounds were subjected to in vitro assays using cellular and biochemical models to validate their interaction with the CB1R and determine their functional impact. Our results reveal FDA-approved compounds that exhibit promising interactions with the CB1R. These findings open up exciting opportunities for DR in various disorders where CB1R signaling is implicated. In conclusion, our integrated computational and experimental approach demonstrates the feasibility of DR for discovering CB1R modulators from existing FDA-approved compounds. By leveraging the wealth of existing pharmacological data, this strategy accelerates the identification of potential therapeutics while reducing development costs and timelines. The findings from this study hold the potential to advance novel treatments for a range of CB1R -associated diseases, presenting a significant step forward in drug discovery research.

The endocannabinoid system (ECS) refers to a complex cell-signaling system highly conserved among species formed by numerous receptors, lipid mediators (endocannabinoids) and synthetic and degradative enzymes. It is widely distributed throughout the body including the CNS, where it participates in synaptic signaling, plasticity and neurodevelopment. Besides, the olfactory ensheathing glia (OEG) present in the olfactory system is also known to play an important role in the promotion of axonal growth and/or myelination. Therefore, both OEG and the ECS promote neurogenesis and oligodendrogenesis in the CNS. Here, we investigated if the ECS is expressed in cultured OEG, by assessing the main markers of the ECS through immunofluorescence, western blotting and qRT-PCR and quantifying the content of endocannabinoids in the conditioned medium of these cells. After that, we investigated whether the production and release of endocannabinoids regulate the differentiation of oligodendrocytes co-cultured with hippocampal neurons, through Sholl analysis in oligodendrocytes expressing O4 and MBP markers. Additionally, we evaluated through western blotting the modulation of downstream pathways such as PI3K/Akt/mTOR and ERK/MAPK, being known to be involved in the proliferation and differentiation of oligodendrocytes and activated by CB1, which is the major endocannabinoid responsive receptor in the brain. Our data show that OEG expresses key genes of the ECS, including the CB1 receptor, FAAH and MAGL. Besides, we were able to identify AEA, 2-AG and AEA related mediators palmitoylethanolamide (PEA) and oleoylethanolamide (OEA), in the conditioned medium of OEG cultures. These cultures were also treated with URB597 10-9 M, a FAAH selective inhibitor, or JZL184 10-9 M, a MAGL selective inhibitor, which led to the increase in the concentrations of OEA and 2-AG in the conditioned medium. Moreover, we found that the addition of OEG conditioned medium (OEGCM) enhanced the complexity of oligodendrocyte process branching in hippocampal mixed cell cultures and that this effect was inhibited by AM251 10-6 M, a CB1 receptor antagonist. However, treatment with the conditioned medium enriched with OEA or 2-AG did not alter the process branching complexity of premyelinating oligodendrocytes, while decreased the branching complexity in mature oligodendrocytes. We also observed no change in the phosphorylation of Akt and ERK 44/42 in any of the conditions used. In conclusion, our data show that the ECS modulates the number and maturation of oligodendrocytes in hippocampal mixed cell cultures.

Many malignant cancers, including breast cancer, have a propensity to invade bones, leading to excruciating bone pain. Opioids are the primary analgesics used to alleviate this cancer-induced bone pain (CIBP) but are associated with numerous severe side effects, including enhanced bone degradation, which significantly impairs patients' quality of life. By contrast, agonists activating only peripheral CB1 receptors (CB1Rs) have been shown to effectively alleviate multiple chronic pain conditions with limited side effects, yet no studies have evaluated their role(s) in CIBP. Here, we demonstrate for the first time that a peripherally selective CB1R agonist can effectively suppress CIBP. Our studies using a syngeneic murine model of CIBP show that both acute and sustained administration of a peripherally restricted CB1R agonist, 4-{2-[-(1E)-1[(4-propylnaphthalen-1-yl)methylidene]-1H-inden-3-yl]ethyl}morpholine (PrNMI), significantly alleviated spontaneous pain behaviors in the animals. This analgesic effect by PrNMI can be reversed by a systemic administration but not spinal injection of SR141716, a selective CB1R antagonist. In addition, the cancer-induced bone loss in the animals was not exacerbated by a repeated administration of PrNMI. Furthermore, catalepsy and hypothermia, the common side effects induced by cannabinoids, were measured at the supratherapeutic doses of PrNMI tested. PrNMI induced mild sedation, yet no anxiety or a decrease in limb movements was detected. Overall, our studies demonstrate that CIBP can be effectively managed by using a peripherally restricted CB1R agonist, PrNMI, without inducing dose-limiting central side effects. Thus, targeting peripheral CB1Rs could be an alternative therapeutic strategy for the treatment of CIBP.

Endocannabinoids (ECBs) depress transmitter release at sites throughout the brain. Here, we describe another form of ECB signaling that triggers a novel form of long-term potentiation (LTP) localized to the lateral perforant path (LPP) which conveys semantic information from cortex to hippocampus. Two cannabinoid CB1 receptor (CB1R) signaling cascades were identified in hippocampus. The first is pregnenolone sensitive, targets vesicular protein Munc18-1 and depresses transmitter release; this cascade is engaged by CB1Rs in Schaffer-Commissural afferents to CA1 but not in the LPP, and it does not contribute to LTP. The second cascade is pregnenolone insensitive and LPP specific; it entails co-operative CB1R/β1-integrin signaling to effect synaptic potentiation via stable enhancement of transmitter release. The latter cascade is engaged during LPP-dependent learning. These results link atypical ECB signaling to the encoding of a fundamental component of episodic memory and suggest a novel route whereby endogenous and exogenous cannabinoids affect cognition.

The aim of this study was to identify and validate the reference genes in cultured human odontoblasts to quantify their cannabinoid receptor transcripts. The most stably transcribed genes in cultured human odontoblast cells were identified using the RefGenes tool and were selected for real-time polymerase chain reaction (PCR) amplification. Human odontoblast cells were differentiated from mesenchymal stem cells using a transforming growth factor-β-supplemented differentiation medium, and total RNA was purified. Reverse transcription-quantitative PCR and relative quantification analyses were performed using the Schefe's method. The relative expression dataset was analyzed to select the most stable genes. The analysis showed that the transcripts of cholinergic receptor nicotinic beta 2 subunit, LIM homeobox transcription factor 1 beta, and family with sequence similarity 223 member B presented the lowest standard deviation (SD) in expression (SD: 0.2, 0.17, and 0.16, respectively). These genes showed similar expression levels as the target genes (cannabinoid receptors). Significant differences were found in the relative expression levels of cannabinoid receptors using the selected genes compared to those calculated using beta actin transcripts as references (p < 0.05). The strategy reported here for searching and verifying new reference genes will aid in the accurate and reliable expression of cannabinoid receptors in human odontoblast cells. [Display omitted] • Cannabinoid receptors play an essential role regulating pain and inflammation. • Reference gene validation is needed to improve the accuracy of RT qPCR results. • Bioinformatic and experimental techniques are suitable to find the most stable gen. • A significant difference in expression is observed normalizing with CRN or β-actin. [ABSTRACT FROM AUTHOR]

Neuroinflammation is a complex biological process that typically originates as a protective response in the brain. This inflammatory process is triggered by the release of pro-inflammatory substances like cytokines, prostaglandins, and reactive oxygen and nitrogen species from stimulated endothelial and glial cells, including those with pro-inflammatory functions, in the outer regions. While neuronal inflammation is common in various central nervous system disorders, the specific inflammatory pathways linked with different immune-mediated cell types and the various factors influencing the blood-brain barrier significantly contribute to disease-specific characteristics. The endocannabinoid system consists of cannabinoid receptors, endogenous cannabinoids, and enzymes responsible for synthesizing and metabolizing endocannabinoids. The primary cannabinoid receptor is CB1, predominantly found in specific brain regions such as the brainstem, cerebellum, hippocampus, and cortex. The presence of CB2 receptors in certain brain components, like cultured cerebellar granular cells, Purkinje fibers, and microglia, as well as in the areas like the cerebral cortex, hippocampus, and cerebellum is also evidenced by immunoblotting assays, radioligand binding, and autoradiography studies. Both CB1 and CB2 cannabinoid receptors exhibit noteworthy physiological responses and possess diverse neuromodulatory capabilities. This review primarily aims to outline the distribution of CB1 and CB2 receptors across different brain regions and explore their potential roles in regulating neuroinflammatory processes.

Background: Mental disorders linked with dysfunction in the temporal cortex, such as anxiety and depression, can increase the morbidity and mortality of people living with HIV (PLWHA). Expressions of both nucleobindin 1 (NUCB1) and cannabinoid receptor 1 (CNR1) in the neurons have been found to alter in patients with depressive disorder, but whether it is involved in the development of depression in the context of HIV infection is unknown. Objectives To investigate the effects of NUCB1 on depressive disorder among PLWHA and preliminarily explore the underlying molecular mechanisms. Methods: Individuals who were newly HIV diagnosed were assessed on the Hospital Anxiety and Depression scale (HADS). Then SHIV-infected rhesus monkeys were used to investigate the possible involvement of the NUCB1 and the CNR1 protein in depression-like behavior. Results: The prevalence rate of depression among PLWHA was 27.33% (41/150). The mechanism results showing elevated NUCB1 levels in cerebrospinal fluid from HIV-infected patients suffering from depression were confirmed compared to those of HIV-infected patients. Moreover, the immunohistochemical analysis indicated the expression of NUCB1 in the temporal cortex neurons of SHIV-infected monkeys was higher than that of the healthy control. Conversely, CNR1 expression was down-regulated at protein levels. Conclusions: Depression symptoms are common among PLWHA and associate with NUCB1 expression increases, and NUCB1 may be a potential target for depression. [ABSTRACT FROM AUTHOR]

Introduction: Cannabidiol (CBD) can be isolated from Cannabis sativa L. or synthetically produced. The aim of this study was to compare the in vitro effects of purified natural and synthetic CBD to establish any pharmacological differences or superiority between sources. Methods: Six purified samples of CBD were obtained, 4 of these were natural and 2 synthetic. The anticancer effects of CBD were assessed in a human ovarian cancer cell line (SKOV-3 cells). The neuroprotective effects of CBD were assessed in human pericytes in a model of stroke (oxygen glucose deprivation [OGD]). The ability of CBD to restore inflammation-induced intestinal permeability was assessed in differentiated human Caco-2 cells (a model of enterocytes). Results: (1) In proliferating and confluent SKOV-3 cells, all CBD samples similarly reduced resazurin metabolism as a marker of cell viability in a concentration-dependent manner (p < 0.001). (2) In pericytes exposed to OGD, all CBD samples similarly reduced cellular damage (measured by lactate dehydrogenase) at 24 h by 31–48% and reduced inflammation (measured by IL-6 secretion) by 30–53%. Attenuation of IL-6 was inhibited by 5HT1A receptor antagonism for all CBD sources. (3) In differentiated Caco-2 cells exposed to inflammation (TNFα and IFNγ, 10 ng/mL for 24 h), each CBD sample increased the speed of recovery of epithelial permeability compared to control (p < 0.05–0.001), which was inhibited by a CB1 receptor antagonist. Conclusion: Our results suggest that there is no pharmacological difference in vitro in the antiproliferative, anti-inflammatory, or permeability effects of purified natural versus synthetic CBD. The purity and reliability of CBD samples, as well as the ultimate pharmaceutical preparation, should all be considered above the starting source of CBD in the development of new CBD medicines.

Endocannabinoid signaling plays crucial roles in human physiology in the function of multiple systems. The two cannabinoid receptors, CB1 and CB2, are cell membrane proteins that interact with both exogenous and endogenous bioactive lipid ligands, or endocannabinoids. Recent evidence has established that endocannabinoid signaling operates within the human kidney, as well as suggests the important role it plays in multiple renal pathologies. CB1, specifically, has been identified as the more prominent ECS receptor within the kidney, allowing us to place emphasis on this receptor. The activity of CB1 has been repeatedly shown to contribute to both diabetic and non-diabetic chronic kidney disease (CKD). Interestingly, recent reports of acute kidney injury (AKI) have been attributed to synthetic cannabinoid use. Therefore, the exploration of the ECS, its receptors, and its ligands can help provide better insight into new methods of treatment for a range of renal diseases. This review explores the endocannabinoid system, with a focus on its impacts within the healthy and diseased kidney.

2‐Monoacylglycerol (2‐MAG) is one of the digestion products of dietary lipids. We recently demonstrated that a 2‐MAG, 2‐arachidonoyl glycerol (2‐AG) potently stimulated cholecystokinin (CCK) secretion via cannabinoid receptor 1 (CB1) in a murine CCK‐producing cell line, STC‐1. CCK plays a crucial role in suppressing postprandial gastric emptying. To examine the effect of 2‐AG on gastric emptying, we performed acetaminophen and phenol red recovery tests under oral or intraperitoneal administration of 2‐AG in mice. Orally administered 2‐AG (25 mg/kg) suppressed the gastric emptying rate in mice, as determined by the acetaminophen absorption test and phenol red recovery test. Intraperitoneal administration of a cholecystokinin A receptor antagonist (0.5 mg/kg) attenuated the gastric inhibitory emptying effect. In addition, both oral (10 mg/kg) and intraperitoneal (0.5 mg/kg) administration of a CB1 antagonist counteracted the 2‐AG‐induced gastric inhibitory effect. Furthermore, intraperitoneal 2‐AG (25 mg/kg) suppressed gastric emptying. These results indicate that 2‐AG exhibits an inhibitory effect on gastric emptying in mice, possibly mediated by stimulating both CCK secretion via CB1 expressed in CCK‐producing cells and acting on CB1 expressed in the peripheral nerves. Our findings provide novel insights into the 2‐MAG‐sensing mechanism in enteroendocrine cells and the physiological role of 2‐MAG. [ABSTRACT FROM AUTHOR]

The anxiolytic and antidepressant properties of cannabidiol (CBD) have been evaluated in several studies. However, the molecular mechanisms involved in these actions remain unclear. A total of 130 male mice were used. CBD's ability to modulate emotional disturbances (anxiety and depressive-like behaviors) was evaluated at different doses in wild-type (CD1; 10, 20 and 30 mg/kg; i.p.) and knockout (CB1KO, CB2KO; GPR55KO; 20 mg/kg) mice. Moreover, CBD effects (20 mg/kg; i.p.) were evaluated in mice previously treated with the CB1r-antagonist SR141716A (2mg/kg; i.p.). Relative gene expression analyses of Cnr1 and Cnr2, Gpr55 and GABA(A)α2 and γ2 receptor subunits were performed in the amygdala (AMY) and hippocampus (HIPP) of CD1 mice. CBD (10 and 20 mg/kg) showed anxiolytic and antidepressant actions in CD1 mice, being more effective at 20 mg/kg. Its administration did not induce anxiolytic actions in CB1KO mice, contrary to CB2KO and GPR55KO. In all of them, the lack of cannabinoid receptors did not modify the antidepressant activity of CBD. Interestingly, the administration of the CB1r antagonist SR141716A blocked the anxiolytic-like activity of CBD. Real-time PCR studies revealed a significant reduction in Cnr1 and GABA(A)α2 and γ2 gene expression in the HIPP and AMY of CD1 mice treated with CBD. Opposite changes were observed in the Cnr2. Indeed, Gpr55 was increased in the AMY and reduced in the HIPP. CB1r appears to play a relevant role in modulating the anxiolytic actions of CBD. Moreover, this study revealed that CBD also modified the gene expression of GABA(A) subunits α2 and γ2 and CB1r, CB2r and GPR55, in a dose- and brain-region-dependent manner, supporting a multimodal mechanism of action for CBD. [ABSTRACT FROM AUTHOR]

Cannabinoid receptor 1 (CB1R), a G protein‐coupled receptor, plays a fundamental role in synaptic plasticity. Abnormal activity and deregulation of CB1R signaling result in a broad spectrum of pathological conditions. CB1R signaling is regulated by receptor desensitization including phosphorylation of residues within the intracellular C terminus by G protein‐coupled receptor kinases (GRKs) that may lead to endocytosis. Furthermore, CB1R signaling is regulated by the protein Src homology 3‐domain growth factor receptor‐bound 2‐like (SGIP1) that hinders receptor internalization, while enhancing CB1R association with β‐arrestin. It has been postulated that phosphorylation of two clusters of serine/threonine residues, 425SMGDS429 and 460TMSVSTDTS468, within the CB1R C‐tail controls dynamics of the association between receptor and its interaction partners involved in desensitization. Several molecular determinants of these events are still not well understood. We hypothesized that the dynamics of these interactions are modulated by SGIP1. Using a panel of CB1Rs mutated in the aforementioned serine and threonine residues, together with an array of Bioluminescence energy transfer‐based (BRET) sensors, we discovered that GRK3 forms complexes with Gβγ subunits of G proteins that largely independent of GRK3's interaction with CB1R. Furthermore, CB1R interacts only with activated GRK3. Interestingly, phosphorylation of two specific residues on CB1R triggers GRK3 dissociation from the desensitized receptor. SGIP1 increases the association of GRK3 with Gβγ subunits of G proteins, and with CB1R. Altogether, our data suggest that the CB1R signalosome complex is dynamically controlled by sequential phosphorylation of the receptor C‐tail and is also modified by SGIP1. [ABSTRACT FROM AUTHOR]

Short tandem repeats (STRs) and variable number of tandem repeats (VNTRs) that have been identified at approximately 0.7 and 0.5 million loci in the human genome, respectively, are highly multi-allelic variations rather than single-nucleotide polymorphisms. The number of repeats of more than a few thousand STRs was associated with the expression of nearby genes, indicating that STRs are influential genetic variations in human traits. Analgesics act on the central nervous system via their intrinsic receptors to produce analgesic effects. In the present study, we focused on STRs and VNTRs in the CNR1 , GRIN2A , PENK , and PDYN genes and analyzed two peripheral pain sensation-related traits and seven analgesia-related traits in postoperative pain management. A total of 192 volunteers who underwent the peripheral pain sensation tests and 139 and 252 patients who underwent open abdominal and orthognathic cosmetic surgeries, respectively, were included in the study. None of the four STRs or VNTRs were associated with peripheral pain sensation. Short tandem repeats in the CNR1 , GRIN2A , and PENK genes were associated with the frequency of fentanyl use, fentanyl dose, and visual analog scale pain scores 3 h after orthognathic cosmetic surgery (Spearman's rank correlation coefficient ρ = 0.199, p = 0.002, ρ = 0.174, p = 0.006, and ρ = 0.135, p = 0.033, respectively), analgesic dose, including epidural analgesics after open abdominal surgery (ρ = −0.200, p = 0.018), and visual analog scale pain scores 24 h after orthognathic cosmetic surgery (ρ = 0.143, p = 0.023), respectively. The associations between STRs in the CNR1 gene and the frequency of fentanyl use and fentanyl dose after orthognathic cosmetic surgery were confirmed by Holm's multiple-testing correction. These findings indicate that STRs in the CNR1 gene influence analgesia in the orofacial region. [ABSTRACT FROM AUTHOR]

Sleep deprivation is commonplace in modern society, Short periods of continuous sleep deprivation (SD) may negatively affect brain and behavioral function and may lead to vehicle accidents and medical errors. Tanshinone IIA (Tan IIA) is an important lipid-soluble component of Salvia miltiorrhiza , which could exert neuroprotective effects. The aim of this study was to investigate the mechanism of neuroprotective effect of Tan IIA on acute sleep deprivation-induced cognitive dysfunction in rats. Tan IIA ameliorated behavioral abnormalities in sleep deprived rats, enhanced behavioral performance in WMW and NOR experiments, increased hippocampal dendritic spine density, and attenuated atrophic loss of hippocampal neurons. Tan IIA enhanced the expression of CB1, PI3K, AKT, STAT3 in rat hippocampus and down-regulated the expression ratio of Bax to Bcl-2. These effects were inhibited by cannabinoid receptor 1 antagonist (AM251). In conclusion, Tan IIA can play a neuroprotective role by activating the CNR1/PI3K/AKT signaling pathway to antagonize apoptosis in the hippocampus and improve sleep deprivation-induced spatial recognition and learning memory dysfunction in rats. Our study suggests that Tan IIA may be a candidate for the prevention of sleep deprivation-induced dysfunction in spatial recognition and learning memory. [ABSTRACT FROM AUTHOR]

Xuelian Zhang,1 Haiqing Zhu,2 Xiaoyan Xing,1 Chunyu Zhang3 1Department of Endocrinology, China-Japan Friendship Hospital, Beijing 100029, People’s Republic of China; 2Department of Endocrinology, Emergency General Hospital, Beijing 100028, People’s Republic of China; 3Department of Statistical Teaching and Research, China-Japan Friendship Hospital, Beijing 100029, People’s Republic of ChinaCorrespondence: Xiaoyan XingDepartment of Endocrinology, China-Japan Friendship Hospital, Chaoyang District, Beijing 100029, People’s Republic of ChinaEmail 3382648198@qq.comBackground: The cannabinoid receptor 1 (CNR1) gene polymorphism is reportedly associated with components of metabolic syndrome and coronary artery diseases in patients with type 2 diabetes mellitus (T2DM). We investigated whether the common variant rs10493353 polymorphism is associated with diabetic nephropathy (DN) in T2DM patients.Patients and Methods: T2DM patients with DN were enrolled as a case group, and patients with only T2DM as a control group. Demographic data and biochemical parameters were collected. The polymerase chain reaction-based restriction fragment length polymorphism technique was used for genotyping. The odds ratio and 90% confidence interval were calculated to assess the association between genotypes and the risk of DN.Results: In total, 320 T2DM patients and 320 DN patients were enrolled. Compared with T2DM patients, the DN patients have a significantly larger body mass index (BMI), longer duration of disease, and higher proportions of smokers, drinkers, and hypertension. The risk of DN was significantly decreased by genotypes AA (OR=0.39, 95% CI=0.23– 0.67) and GA (OR=0.53, 95% CI=0.37– 0.75) vs GG (codominant model), GA/AA vs GG (OR=0.49, 95% CI=0.35– 0.67; dominant model), AA vs GG/GA (OR=0.47, 95% CI=0.28– 0.80; recessive model), and the A allele (OR=0.52, 95% CI=0.40– 0.68; allele model). Multiple logistic regressions still show significant levels. Negative interactions were found between gene and clinical parameters, including drinking, smoking, BMI, and hypertension.Conclusion: The A allele of CNR1 gene rs10493353 may be a protective factor for DN in T2DM patients. The risk factors of DN can affect the protective role of A allele in the progression of DN.Keywords: diabetes mellitus, diabetic nephropathy, gene polymorphism, cannabinoid receptor 1

Nod-like receptor (NLR) family pyrin domain containing 3 (NLRP3) has been regarded as an important initiator or promoter in multiple inflammatory diseases. However, the relationship between cannabinoid receptor 1 (CB1) and macrophage NLRP3 inflammasome and the corresponding molecular mechanism in liver inflammation remain unclear. Mouse liver injury models were induced by carbon tetrachloride (CCl4) or methionine-choline-deficient and high fat (MCDHF) diet. Human liver tissues were obtained from patients with different chronic liver diseases. CB1 expression was increased in liver tissue and macrophages of CCl4- and MCDHF-treated mice, positively correlated with NLRP3. CB1 agonist ACEA (Arachiodonyl-2’-Chloroethylamide) promoted NLRP3 expression and NLRP3 inflammasome activation in macrophages. CB1 blockade with its antagonist AM281 reduced NLRP3 expression, inflammasome activation, and liver inflammation in CCl4- and MCDHF-treated mice. MicroRNA-30b-5p (miR-30b-5p), screened by the intersection of bioinformatics databases and downregulated miRNAs in injured liver, negatively correlated with NLRP3 in mouse and human liver. miR-30b-5p was involved in CB1-mediated activation of NLRP3 inflammasome in macrophages by directly targeting NLRP3. Importantly, administration of miR-30b-5p agomir targeted NLRP3 and attenuated liver inflammation in the injured liver. Altogether, CB1/miR-30b-5p axis modulates NLRP3 expression and NLPR3 inflammasome activation in macrophages during liver inflammation, which provides a potential target for liver disease.

Short tandem repeats (STRs) and variable number of tandem repeats (VNTRs) that have been identified at approximately 0.7 and 0.5 million loci in the human genome, respectively, are highly multi-allelic variations rather than single-nucleotide polymorphisms. The number of repeats of more than a few thousand STRs was associated with the expression of nearby genes, indicating that STRs are influential genetic variations in human traits. Analgesics act on the central nervous system via their intrinsic receptors to produce analgesic effects. In the present study, we focused on STRs and VNTRs in the CNR1, GRIN2A, PENK, and PDYN genes and analyzed two peripheral pain sensation-related traits and seven analgesia-related traits in postoperative pain management. A total of 192 volunteers who underwent the peripheral pain sensation tests and 139 and 252 patients who underwent open abdominal and orthognathic cosmetic surgeries, respectively, were included in the study. None of the four STRs or VNTRs were associated with peripheral pain sensation. Short tandem repeats in the CNR1, GRIN2A, and PENK genes were associated with the frequency of fentanyl use, fentanyl dose, and visual analog scale pain scores 3 h after orthognathic cosmetic surgery (Spearman’s rank correlation coefficient ρ = 0.199, p = 0.002, ρ = 0.174, p = 0.006, and ρ = 0.135, p = 0.033, respectively), analgesic dose, including epidural analgesics after open abdominal surgery (ρ = −0.200, p = 0.018), and visual analog scale pain scores 24 h after orthognathic cosmetic surgery (ρ = 0.143, p = 0.023), respectively. The associations between STRs in the CNR1 gene and the frequency of fentanyl use and fentanyl dose after orthognathic cosmetic surgery were confirmed by Holm’s multiple-testing correction. These findings indicate that STRs in the CNR1 gene influence analgesia in the orofacial region.

Sleep deprivation is commonplace in modern society, Short periods of continuous sleep deprivation (SD) may negatively affect brain and behavioral function and may lead to vehicle accidents and medical errors. Tanshinone IIA (Tan IIA) is an important lipid-soluble component of Salvia miltiorrhiza, which could exert neuroprotective effects. The aim of this study was to investigate the mechanism of neuroprotective effect of Tan IIA on acute sleep deprivation-induced cognitive dysfunction in rats. Tan IIA ameliorated behavioral abnormalities in sleep deprived rats, enhanced behavioral performance in WMW and NOR experiments, increased hippocampal dendritic spine density, and attenuated atrophic loss of hippocampal neurons. Tan IIA enhanced the expression of CB1, PI3K, AKT, STAT3 in rat hippocampus and down-regulated the expression ratio of Bax to Bcl-2. These effects were inhibited by cannabinoid receptor 1 antagonist (AM251). In conclusion, Tan IIA can play a neuroprotective role by activating the CNR1/PI3K/AKT signaling pathway to antagonize apoptosis in the hippocampus and improve sleep deprivation-induced spatial recognition and learning memory dysfunction in rats. Our study suggests that Tan IIA may be a candidate for the prevention of sleep deprivation-induced dysfunction in spatial recognition and learning memory.

Cholecystokinin (CCK) is a peptide hormone secreted from enteroendocrine cells and regulates the exocrine pancreas, gastric motility, and appetite. Dietary triacylglycerols are hydrolyzed to fatty acids (FA) and 2‐monoacylglycerols (2‐MAG) in the small intestine. Although it is well known that FA stimulate CCK secretion, whether 2‐MAG have the CCK‐releasing activity remains unclear. We examined the CCK‐releasing activity of four commercially available 2‐MAG in a murine CCK‐producing cell line, STC‐1, and the molecular mechanism underlying 2‐MAG‐induced CCK secretion. CCK released from the cells was measured using ELISA. Among four 2‐MAG (2‐palmitoyl, 2‐oleoyl, 2‐linoleoyl, and 2‐arachidonoyl monoacylglycerols) examined, 2‐arachidonoyl glycerol (2‐AG) potently stimulated CCK secretion in a dose‐dependent manner. Structurally related compounds, such as 2‐arachidonoyl glycerol ether and 1‐arachidonoyl glycerol, did not stimulate CCK secretion. Both arachidonic acid and 2‐AG stimulated CCK secretion at 100 μM, but only 2‐AG did at 50 μM. 2‐AG‐induced CCK secretion but not arachidonic acid‐induced CCK secretion was attenuated by treatment with a cannabinoid receptor 1 (CB1) antagonist. These results indicate that a specific 2‐MAG, 2‐AG, directly stimulates CCK secretion via CB1. [ABSTRACT FROM AUTHOR]

Post-traumatic stress disorder (PTSD) develops in a subset of individuals exposed to a trauma with core features being increased anxiety and impaired fear extinction. To model the heterogeneity of PTSD behavioral responses, we exposed male Sprague–Dawley rats to predator scent stress once for 10 min and then assessed anxiety-like behavior 7 days later using the elevated plus maze and acoustic startle response. Rats displaying anxiety-like behavior in both tasks were classified as stress Susceptible, and rats exhibiting behavior no different from un-exposed Controls were classified as stress Resilient. In Resilient rats, we previously found increased mRNA expression of mGlu5 in the amygdala and prefrontal cortex (PFC) and CB1 in the amygdala. Here, we performed fluorescent in situ hybridization (FISH) to determine the subregion and cell-type-specific expression of these genes in Resilient rats 3 weeks after TMT exposure. Resilient rats displayed increased mGlu5 mRNA expression in the basolateral amygdala (BLA) and the infralimbic and prelimbic regions of the PFC and increased BLA CB1 mRNA. These increases were limited to glutamatergic cells. To test the necessity of mGlu5 for attenuating TMT-conditioned contextual fear 3 weeks after TMT conditioning, intra-BLA infusions of the mGlu5 negative allosteric modulator MTEP were administered prior to context re-exposure. In TMT-exposed Resilient rats, but not Controls, MTEP increased freezing on the day of administration, which extinguished over two additional un-drugged sessions. These results suggest that increased mGlu5 expression in BLA glutamate neurons contributes to the behavioral flexibility observed in stress-Resilient animals by facilitating a capacity for extinguishing contextual fear associations. [ABSTRACT FROM AUTHOR]