anandamide and Multiple-Sclerosis

Recent years have yielded significant advances in our understanding of microglia, the immune cells of the central nervous system (CNS). Microglia are key players in CNS development, immune surveillance, and the maintenance of proper neuronal function throughout life. In the healthy brain, homeostatic microglia have a unique molecular signature. In neurological diseases, microglia become activated and adopt distinct transcriptomic signatures, including disease-associated microglia (DAM) implicated in neurodegenerative disorders. Homeostatic microglia synthesise the endogenous cannabinoids 2-arachidonoylglycerol and anandamide and express the cannabinoid receptors CB1 and CB2 at constitutively low levels. Upon activation, microglia significantly increase their synthesis of endocannabinoids and upregulate their expression of CB2 receptors, which promote a protective microglial phenotype by enhancing their production of neuroprotective factors and reducing their production of pro-inflammatory factors. Here, we summarise the effects of the microglial cannabinoid system in the CNS demyelinating disease multiple sclerosis, the neurodegenerative diseases Alzheimer's disease, Parkinson's disease and amyotrophic lateral sclerosis, chronic inflammatory and neuropathic pain, and psychiatric disorders including depression, anxiety and schizophrenia. We discuss the therapeutic potential of cannabinoids in regulating microglial activity and highlight the need to further investigate their specific microglia-dependent immunomodulatory effects.

Topics: Alzheimer Disease; Amyotrophic Lateral Sclerosis; Anxiety Disorders; Arachidonic Acids; Chronic Pain; Depressive Disorder; Endocannabinoids; Glycerides; Humans; Mental Disorders; Microglia; Multiple Sclerosis; Neuralgia; Neurodegenerative Diseases; Parkinson Disease; Polyunsaturated Alkamides; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2; Schizophrenia

The discovery of cannabinoid receptors at the beginning of the 1990s, CB1 cloned in 1990 and CB2 cloned in 1993, and the availability of selective and potent cannabimimetics could only be justified by the existence of endogenous ligands that are capable of binding to them. Thus, the characterisation and cloning of the first cannabinoid receptor (CB1) led to the isolation and characterisation of the first endocannabinoid, arachidonoylethanolamide (AEA), two years later and the subsequent identification of a family of lipid transmitters known as the fatty acid ester 2-arachidonoylglycerol (2-AG). The endogenous cannabinoid system is a complex signalling system that comprises transmembrane endocannabinoid receptors, their endogenous ligands (the endocannabinoids), the specific uptake mechanisms and the enzymatic systems related to their biosynthesis and degradation. The endocannabinoid system has been implicated in a wide diversity of biological processes, in both the central and peripheral nervous systems, including memory, learning, neuronal development, stress and emotions, food intake, energy regulation, peripheral metabolism, and the regulation of hormonal balance through the endocrine system. In this context, this article will review the current knowledge of the therapeutic potential of cannabinoid receptor as a target in Alzheimer's disease and other less well-known diseases that include, among others, multiple sclerosis, bone metabolism, and Fragile X syndrome. The therapeutic applications will be addressed through the study of cannabinoid agonists acting as single drugs and multi-target drugs highlighting the CB2 receptor agonist.

Topics: Alzheimer Disease; Animals; Arachidonic Acids; Autism Spectrum Disorder; Bone Diseases; Cannabinoid Receptor Agonists; Cardiovascular Diseases; Drug Discovery; Endocannabinoids; Fragile X Syndrome; Humans; Molecular Targeted Therapy; Multiple Sclerosis; Polyunsaturated Alkamides; Receptor, Cannabinoid, CB2; Receptors, Cannabinoid

Over the last decade, the endocannabinoid system (ECS) has emerged as a potential target for multiple sclerosis (MS) management. A growing amount of evidence suggests that cannabinoids may be neuroprotective during CNS inflammation. Advances in the understanding of the physiology and pharmacology of the ECS have potentiated the interest of several components of this system as useful biological targets for disease management. Alterations of the ECS have been recently implicated in a number of neuroinflammatory and neurodegenerative conditions, so that the pharmacological modulation of cannabinoid (CB) receptors and/or of the enzymes controlling synthesis, transport, and degradation of these lipid mediators is considered an option to treat several neurological diseases. This chapter focuses on our current understanding of the function of anandamide (AEA), its biological and therapeutic implications, as well as a description of its effects on neuroimmune modulation.

Topics: Animals; Arachidonic Acids; Cannabinoid Receptor Modulators; Cytokines; Endocannabinoids; Humans; Immunologic Factors; Interleukin-12; Multiple Sclerosis; Neuroglia; Neuroprotective Agents; Polyunsaturated Alkamides; Signal Transduction

Bayer AG has recently announced that it acquired exclusive rights for the marketing of GW Pharmaceuticals' new medicine Sativex in Europe and in other regions. Sativex is a sublingual spray on Cannabis extract basis, and is equipped with an electronic tool to facilitate accurate dosing and to prevent misuses. It is standardized for the THC and CBD. The new analgesic is proposed for the treatment of muscle spasticity and pains accompanying multiple sclerosis and as an efficient analgetic for neurogenic pain not responding well to opioids and to other therapies available. The entirely new mechanism of action through the recently discovered cannabinoid receptor system may offer a real therapeutic potential to the drug. Although the Government of Netherlands has authorized the sale of pharmaceutical grade Cannabis herb by pharmacies in the Netherlands, the availability on the pharmaceutical market of the registered preparation may render requests for the authorization of the smoking of Cannabis herb (marihuana) by individuals suffering of multiple sclerosis, neurogenic pain, AIDS wasting syndrome unnecessary. Nevertheless, the "old chameleon" plant Cannabis appears to gradually regain its previous status in mainstream therapy and pharmacy. As long as the plant Cannabis and its products continue to be classified as narcotic drugs, medical use of the new preparation will need close supervision.

Topics: Analgesics; Arachidonic Acids; Cannabidiol; Cannabinoids; Cannabis; Dronabinol; Drug Combinations; Drugs, Investigational; Endocannabinoids; Europe; Humans; Multiple Sclerosis; Pain; Plant Extracts; Polyunsaturated Alkamides

Endocannabinoids are a new class of lipids, which include amides, esters and ethers of long chain polyunsaturated fatty acids. Anandamide (N-arachidonoylethanolamine; AEA) and 2-arachidonoylglycerol are the main endogenous agonists of cannabinoid receptors, able to mimic several pharmacological effects of Delta(9)-tetrahydrocannabinol, the active principle of Cannabis sativa preparations like hashish and marijuana. It is known that the activity of AEA is limited by cellular uptake through a specific membrane transporter, followed by intracellular degradation by a fatty acid amide hydrolase. Together with AEA and congeners these proteins form the "endocannabinoid system". The endogenous cannabinoids were identified in brain, and also in neuronal and endothelial cells, suggesting a potential role as modulators in the central nervous system and in the periphery. This review summarises the metabolic routes for the synthesis and degradation of AEA, and the latest advances in the involvement of this lipid in neurovascular biology. In addition, the therapeutic potential of the modulation of endocannabinoid metabolism for neuronal and vascular system will be also reviewed.

Topics: Animals; Arachidonic Acids; Blood Vessels; Cannabinoid Receptor Modulators; Cardiovascular System; Central Nervous System; Endocannabinoids; Humans; Multiple Sclerosis; Nervous System; Neurodegenerative Diseases; Polyunsaturated Alkamides

The endocannabinoid system (ECS) is involved in the pathophysiology of multiple sclerosis (MS), and relief from pain and spasticity has been reported in MS patients self-medicating with marijuana. A cannabis-based medication containing Delta(9)-tetrahydrocannabinol and cannabidiol (Sativex) has been approved in some countries for the treatment of MS-associated pain. The effects of this pharmaceutical preparation on other clinically relevant aspects of MS pathophysiology, however, are still unclear. In 20 MS patients, we measured the effects of Sativex on clinically measured spasticity and on neurophysiological and laboratory parameters that correlate with spasticity severity or with the modulation of the ECS. Sativex failed to affect spasticity and stretch reflex excitability. This compound also failed to affect the synthesis and the degradation of the endocannabinoid anandamide, as well as the expression of both CB1 and CB2 cannabinoid receptors in various subpopulations of peripheral lymphocytes.

Topics: Adult; Arachidonic Acids; Cannabidiol; Cannabinoids; Dronabinol; Drug Combinations; Endocannabinoids; Female; Humans; Lymphocytes; Male; Middle Aged; Multiple Sclerosis; Muscle Spasticity; Pain; Plant Extracts; Polyunsaturated Alkamides; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2; Reflex, Stretch; Treatment Outcome; Young Adult

A combination of molecular modeling and structure-activity relationship studies has been used to fine-tune CB2 selectivity in the chromenopyrazole ring, a versatile CB1/CB2 cannabinoid scaffold. Thus, a series of 36 new derivatives covering a wide range of structural diversity has been synthesized, and docking studies have been performed for some of them. Biological evaluation of the new compounds includes, among others, cannabinoid binding assays, functional studies, and surface plasmon resonance measurements. The most promising compound [43 (PM226)], a selective and potent CB2 agonist isoxazole derivative, was tested in the acute phase of Theiler's murine encephalomyelitis virus-induced demyelinating disease (TMEV-IDD), a well-established animal model of primary progressive multiple sclerosis. Compound 43 dampened neuroinflammation by reducing microglial activation in the TMEV.

Topics: Dose-Response Relationship, Drug; HEK293 Cells; Humans; Models, Molecular; Molecular Structure; Multiple Sclerosis; Pyrazoles; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2; Structure-Activity Relationship

Monocytes are believed to be involved in the immunopathogenesis of multiple sclerosis (MS). The aim of this study was to investigate their role in MS and their immunomodulation by the endocannabinoid system (ECS), a novel target for the treatment of this disease. We compared the level of cytokine production from monocytes in healthy subjects and MS patients upon stimulation with viral or bacterial Toll-like receptors (TLR) and we evaluated the ECS immunomodulatory role in these cells. Here we show that MS monocytes produced more TNF-α, IL-12 and IL-6 following activation of TLR2/4 with LPS or of TLR5 with flagellin, as opposed to TLR7/8 stimulation with R848. Furthermore AEA, the main endocannabinoid, suppressed cytokine production and release from healthy monocytes upon stimulation with both bacterial and viral TLR receptors but not in cells from MS patients, where its immunosuppressive activity was TLR7/8-dependent. Altered expression levels of key ECS members in MS monocytes paralleled these data. Our data disclose a distinct immunomodulatory effect of AEA and an alteration of AEA-related members of the ECS in monocytes from MS patients that involves viral but not bacterial TLR. These findings not only may help to better understand the role of monocytes in MS immunopathogenesis but also could be of help to exploit new endocannabinoid-based drugs that target innate immune cells.

Topics: Adult; Arachidonic Acids; Endocannabinoids; Female; Humans; Immunologic Factors; Immunosuppressive Agents; Interleukin-12; Interleukin-6; Lipids; Male; Monocytes; Multiple Sclerosis; Polyunsaturated Alkamides; Signal Transduction; Toll-Like Receptor 2; Toll-Like Receptor 4; Tumor Necrosis Factor-alpha

Cannabinoids affect immune responses in ways that may be beneficial for autoimmune diseases. We sought to determine whether chronic Cannabis use differentially modulates a select number of immune parameters in healthy controls and individuals with multiple sclerosis (MS cases). Subjects were enrolled and consented to a single blood draw, matched for age and BMI. We measured monocyte migration isolated from each subject, as well as plasma levels of endocannabinoids and cytokines. Cases met definition of MS by international diagnostic criteria. Monocyte cell migration measured in control subjects and individuals with MS was similarly inhibited by a set ratio of phytocannabinoids. The plasma levels of CCL2 and IL17 were reduced in non-naïve cannabis users irrespective of the cohorts. We detected a significant increase in the endocannabinoid arachidonoylethanolamine (AEA) in serum from individuals with MS compared to control subjects, and no significant difference in levels of other endocannabinoids and signaling lipids irrespective of Cannabis use. Chronic Cannabis use may affect the immune response to similar extent in individuals with MS and control subjects through the ability of phytocannabinoids to reduce both monocyte migration and cytokine levels in serum. From a panel of signaling lipids, only the levels of AEA are increased in individuals with MS, irrespective of Cannabis use or not. Our results suggest that both MS cases and controls respond similarly to chronic Cannabis use with respect to the immune parameters measured in this study.

Topics: Adult; Arachidonic Acids; Cannabinoids; Cannabis; Case-Control Studies; Cell Movement; Chemokine CCL2; Cross-Sectional Studies; Endocannabinoids; Female; Humans; Interleukin-17; Male; Marijuana Smoking; Monocytes; Multiple Sclerosis; Polyunsaturated Alkamides

The immunopathogenesis of multiple sclerosis (MS) has always been thought to be driven by chronically activated and autoreactive Th-1 and Th-17 cells. Recently, dendritic cells (DCs) have also been thought to significantly contribute to antigenic spread and to maturation of adaptive immunity, and have been linked with disease progression and exacerbation. However, the role of DCs in MS pathogenesis remains poorly understood.. We compared the level of cytokine production by myeloid DCs (mDCs) and plasmacytoid DCs (pDCs) in healthy subjects and MS patients, following in vitro stimulation of Toll-like receptors 7/8. We also evaluated the effect of the main endocannabinoid, anandamide (AEA), in these DC subsets and correlated cytokine levels with defects in the endocannabinoid system.. mDCs obtained from MS patients produce higher levels of interleukin-12 and interleukin-6, whereas pDCs account for lower levels of interferon-α compared to healthy subjects. AEA significantly inhibited cytokine production from healthy mDCs and pDCs, as well as their ability to induce Th-1 and Th-17 lineages. Moreover, we found that in MS only pDCs lack responsiveness to cytokine inhibition induced by AEA. Consistently, this specific cell subset expresses higher levels of the anandamide hydrolase fatty acid amide hydrolase (FAAH).. Our data disclose a distinct immunomodulatory effect of AEA in mDCs and pDCs from MS patients, which may reflect an alteration of the expression of FAAH, thus forming the basis for the rational design of new endocannabinoid-based immunotherapeutic agents targeting a specific cell subset.

Topics: Adolescent; Adult; Aged; Amidohydrolases; Arachidonic Acids; Cannabinoid Receptor Agonists; Cell Differentiation; Cells, Cultured; Cytokines; Dendritic Cells; Endocannabinoids; Female; Gene Expression Regulation; Humans; Male; Middle Aged; Multiple Sclerosis; Myeloid Cells; Polyunsaturated Alkamides; Receptor, Cannabinoid, CB2; RNA, Messenger; Toll-Like Receptor 7; Toll-Like Receptor 8; Young Adult

Theiler's virus (TMEV) infection of the central nervous system (CNS) induces an immune-mediated demyelinating disease in susceptible mouse strains and serves as a relevant infection model for human multiple sclerosis (MS). The endocannabinoid system represents a novel therapeutic target for autoimmune and chronic inflammatory diseases due to its anti-inflammatory properties by regulating cytokine network. IL-12p70 and IL-23 are functionally related heterodimeric cytokines that play a crucial role in the pathogenesis of MS. In the present study we showed that the endocannabinoid anandamide (AEA) downregulated the gene expression of IL-12p70 and IL-23 forming subunits mRNAs in the spinal cord of TMEV-infected mice and ameliorated motor disturbances. This was accompanied by significant decreases on the serological levels of IL-12p70/IL-23 and more interestingly, of IL-17A. In contrast, serum levels of IL-10 resulted elevated. In addition, we studied the signalling pathways involved in the regulation of IL-12p70/IL-23 and IL-10 expression in TMEV-infected microglia and addressed the possible interactions of AEA with these pathways. AEA acted through the ERK1/2 and JNK pathways to downregulate IL-12p70 and IL-23 while upregulating IL-10. These effects were partially mediated by CB2 receptor activation. We also described an autocrine circuit of cross-talk between IL-12p70/IL-23 and IL-10, since endogenously produced IL-10 negatively regulates IL-12p70 and IL-23 cytokines in TMEV-infected microglia. This suggests that by altering the cytokine network, AEA could indirectly modify the type of immune responses within the CNS. Accordingly, pharmacological modulation of endocannabinoids might be a useful tool for treating neuroinflammatory diseases.

Topics: Adaptive Immunity; Analysis of Variance; Animals; Arachidonic Acids; Cannabinoid Receptor Modulators; Cardiovirus Infections; Disease Models, Animal; Down-Regulation; Endocannabinoids; Female; Interleukin-10; Interleukin-12; Interleukin-23; Interleukins; Mice; Microglia; Multiple Sclerosis; Nervous System Autoimmune Disease, Experimental; Neuroimmunomodulation; Polyunsaturated Alkamides; Protein Subunits; Receptor Cross-Talk; RNA, Messenger; Signal Transduction; Statistics, Nonparametric; Theilovirus

We provide evidence that TWIK-related acid-sensitive potassium channel 1 (TASK1), a member of the family of two-pore domain potassium channels relevant for setting the resting membrane potential and balancing neuronal excitability that is expressed on T cells and neurons, is a key modulator of T cell immunity and neurodegeneration in autoimmune central nervous system inflammation. After induction of experimental autoimmune encephalomyelitis, an experimental model mimicking multiple sclerosis, TASK1(-/-) mice showed a significantly reduced clinical severity and markedly reduced axonal degeneration compared with wild-type controls. T cells from TASK1(-/-) mice displayed impaired T cell proliferation and cytokine production, while the immune repertoire is otherwise normal. In addition to these effects on systemic T cell responses, TASK1 exhibits an independent neuroprotective effect which was demonstrated using both a model of acutely prepared brain slices cocultured with activated T cells as well as in vitro cultivation experiments with isolated optic nerves. Anandamide, an endogenous cannabinoid and inhibitor of TASK channels, reduced outward currents and inhibited effector functions of T cells (IFN-gamma production and proliferation); an effect completely abrogated in TASK1(-/-) mice. Accordingly, preventive blockade of TASK1 significantly ameliorated experimental autoimmune encephalomyelitis after immunization. Therapeutic application of anandamide significantly reduced disease severity and was capable of lowering progressive loss of brain parenchymal volume as assessed by magnetic resonance imaging. These data support the identification and characterization of TASK1 as potential molecular target for the therapy of inflammatory and degenerative central nervous system disorders.

Topics: Animals; Arachidonic Acids; Axons; Cells, Cultured; Coculture Techniques; Disease Progression; Encephalomyelitis, Autoimmune, Experimental; Endocannabinoids; Female; Immunophenotyping; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; Multiple Sclerosis; Nerve Degeneration; Nerve Tissue Proteins; Polyunsaturated Alkamides; Potassium Channel Blockers; Potassium Channels, Tandem Pore Domain; Spinal Cord; Spleen; T-Lymphocyte Subsets; Tissue Culture Techniques

Multiple sclerosis (MS) is a chronic inflammatory disease of the CNS. Therapies that affect the endocannabinoid (EC) system may have immunomodulatory, symptomatic and neuroprotective effects.. The aim of this study was to determine how levels of EC and related compounds are altered in MS.. Plasma and whole blood were collected from 24 MS patients (10 relapsing-remitting (RR); 8 secondary-progressive (SP); 6 primary-progressive (PP); 19 females; 25-66 years) and 17 controls (10 females; 22-62 years). Plasma EC and related compounds were quantified by liquid chromatography-tandem mass spectrometry. Fatty acid amide hydrolase (FAAH), cannabinoid receptors CB(1) and CB(2) mRNA were measured by quantitative reverse transcriptase-polymerase chain reaction.. Anandamide (AEA) and palmitoylethanolamide (PEA) were higher in RRMS compared to controls (p=0.001 and p=0.027). AEA, PEA and oleoylethanolamide were also increased in SPMS plasma (p=0.001, p=0.004, and p=0.005). PPMS patients had higher AEA plasma levels compared to controls (p=0.009). FAAH mRNA was decreased in SPMS (p=0.04) but not in RRMS or PPMS blood. CB(1) (p=0.012) and CB(2) mRNA (p=0.003) were increased in the PPMS.. The EC system is altered in MS. It may be dynamically modulated depending on the subtype of the disease, but further studies with larger subgroups are needed to confirm this.

Topics: Adult; Aged; Amides; Amidohydrolases; Arachidonic Acids; Brain; Brain Chemistry; Cannabinoid Receptor Modulators; Cannabinoids; Chromatography, Liquid; Cytoprotection; Disability Evaluation; Endocannabinoids; Ethanolamines; Female; Humans; Male; Mass Spectrometry; Middle Aged; Multiple Sclerosis; Oleic Acids; Palmitic Acids; Polyunsaturated Alkamides; Receptors, Cannabinoid; RNA, Messenger

Endocannabinoids (eCBs) play a role in the modulation of neuroinflammation, and experimental findings suggest that they may be directly involved in the pathogenesis of multiple sclerosis (MS). The objective of our study was to measure eCB levels in the cerebrospinal fluid (CSF) of patients with MS.. Arachidonoylethanolamine (anandamide, AEA), palmotylethanolamide (PEA), 2-arachidonoylglycerol (2-AG) and oleoylethanolamide (OEA) levels were measured in the CSF of 50 patients with MS and 20 control subjects by isotope dilution gas-chromatography/mass-spectrometry. Patients included 35 patients with MS in the relapsing-remitting (RR) form of the disease, 20 in a stable clinical phase and 15 during a relapse, and 15 patients with MS in the secondary progressive (SP) form.. Significantly reduced levels of all the tested eCBs were found in the CSF of patients with MS compared to control subjects, with lower values detected in the SP MS group. Higher levels of AEA and PEA, although below those of controls, were found in the CSF of RR MS patients during a relapse. Higher levels of AEA, 2-AG and OEA were found in patients with MRI gadolinium-enhancing (Gd+) lesions.. The present findings suggest the presence of an impaired eCB system in MS. Increased CSF levels of AEA during relapses or in RR patients with Gd+ lesions suggest its potential role in limiting the ongoing inflammatory process with potential neuroprotective implications. These findings provide further support for the development of drugs targeting eCBs as a potential pharmacological strategy to reduce the symptoms and slow disease progression in MS.

Topics: Adult; Arachidonic Acids; Brain; Cannabinoid Receptor Modulators; Disability Evaluation; Disease Progression; Endocannabinoids; Female; Gas Chromatography-Mass Spectrometry; Glycerides; Humans; Inflammation; Magnetic Resonance Imaging; Male; Multiple Sclerosis; Oleic Acids; Polyunsaturated Alkamides; Severity of Illness Index

The ability of cannabinoids to modulate both inflammatory and degenerative neuronal damage prompted investigations on the potential benefits of such compounds in multiple sclerosis (MS) and in animal models of this disorder. Here we measured endocannabinoid levels, metabolism and binding, and physiological activities in 26 patients with MS (17 females, aged 19-43 years), 25 healthy controls and in mice with experimental autoimmune encephalomyelitis (EAE), a preclinical model of MS. Our results show that MS and EAE are associated with significant alterations of the endocannabinoid system. We found that anandamide (AEA), but not 2-arachidonoylglycerol (2-AG), was increased in the CSF of relapsing MS patients. AEA concentrations were also higher in peripheral lymphocytes of these patients, an effect associated with increased synthesis and reduced degradation of this endocannabinoid. Increased synthesis, reduced degradation, and increased levels of AEA were also detected in the brains of EAE mice in the acute phase of the disease, possibly accounting for its anti-excitotoxic action in this disorder. Accordingly, neurophysiological recordings from single neurons confirmed that excitatory transmission in EAE slices is inhibited by CB1 receptor activation, while inhibitory transmission is not. Our study suggests that targeting the endocannabinoid system might be useful for the treatment of MS.

Topics: Acute Disease; Adult; Animals; Arachidonic Acids; Brain; Cannabinoid Receptor Modulators; Corpus Striatum; Disease Models, Animal; Dronabinol; Electrophysiology; Encephalomyelitis, Autoimmune, Experimental; Endocannabinoids; Female; Glycerides; Humans; Lymphocytes; Magnetic Resonance Imaging; Male; Mice; Mice, Inbred C57BL; Multiple Sclerosis; Neuroprotective Agents; Patch-Clamp Techniques; Polyunsaturated Alkamides; Synaptic Transmission; Tissue Culture Techniques

To date, UCM707, (5Z,8Z,11Z,14Z)-N-(3-furylmethyl)eicosa-5,8,11,14-tetraenamide, has the highest potency and selectivity in vitro and in vivo as inhibitor of the endocannabinoid uptake. This may enable this compound to potentiate endocannabinoid transmission, with minimal side effects, in the treatment of several neurological disorders. In the present study, we examined whether the treatment with UCM707 produced beneficial effects, as other cannabinoid-related compounds have already shown, to alleviate motor deterioration or to delay/arrest neurodegeneration, in several models of neurological diseases such as Huntington's disease (HD), Parkinson's disease (PD) and multiple sclerosis (MS). UCM707 exhibited a notable anti-hyperkinetic activity in a rat model of HD generated by bilateral intrastriatal application of 3-nitropropionic acid. This effect was possibly associated with an amelioration of GABA and glutamate deficits induced by the toxin in the globus pallidus and the substantia nigra, respectively. However, UCM707 did not protect against the death of GABAergic neurons that occurs in rats with striatal atrophy generated by unilateral application of malonate, another animal model of HD, which is more useful to test neuroprotective strategies. In addition, UCM707 did not provide neuroprotection in rats with unilateral lesions of the nigrostriatal dopaminergic neurons caused by 6-hydroxydopamine, a rat model of PD. This was possibly due to the fact that UCM707 is devoid of anti-oxidant properties since another uptake inhibitor, AM404, that has these properties acted as a protective agent. Lastly, UCM707 was also unable to inhibit the development of the neurological impairment of rats with experimental autoimmune encephalomyelitis (EAE), an acute model of MS. However, UCM707, like other endocannabinoid uptake inhibitors reported previously, significantly reduced spasticity of the hindlimbs in a chronic relapsing EAE mice, a chronic model of MS. In summary, UCM707 might be a promising compound in HD to alleviate motor symptoms, which represents an important goal considering the current lack of efficient pharmacological treatments in this basal ganglia disorder. However, the compound was unable to delay neurodegeneration in this disorder and also in PD. In addition, UCM707 did not produce any neurological recovery from inflammatory attack in an EAE rat model of MS, although it retained the classic anti-spastic action shown by other uptake inhibitors in the

Topics: 3,4-Dihydroxyphenylacetic Acid; Analysis of Variance; Animals; Arachidonic Acids; Brain Chemistry; Disease Models, Animal; Disease Progression; Dopamine; Dose-Response Relationship, Drug; Drug Interactions; Electrochemistry; Encephalomyelitis, Autoimmune, Experimental; Endocannabinoids; Exploratory Behavior; Furans; gamma-Aminobutyric Acid; Glutamate Decarboxylase; Glutamic Acid; Huntington Disease; Male; Malonates; Movement Disorders; Multiple Sclerosis; Nitro Compounds; Oxidopamine; Polyunsaturated Alkamides; Propionates; Rats; Rats, Sprague-Dawley; Time Factors; Tyrosine 3-Monooxygenase

We previously reported that the compound O-2093 is a selective inhibitor of the reuptake of the endocannabinoid anandamide (AEA). We have now re-examined the activity of O-2093 in vivo and synthesized four structural analogs (O-2247, O-2248, O-3246, and O-3262), whose activity was assessed in: (a) binding assays carried out with membranes from cells overexpressing the human CB(1) and CB(2) receptors; (b) assays of transient receptor potential of the vanilloid type-1 (TRPV1) channel functional activity (measurement of [Ca(2+)](i)); (c) [(14)C]AEA cellular uptake and hydrolysis assays in rat basophilic leukaemia (RBL-2H3) cells; (d) the mouse 'tetrad' tests (analgesia on a hot plate, immobility on a 'ring', rectal hypothermia and hypolocomotion in an open field); and (e) the limb spasticity test in chronic relapsing experimental allergic encephalomyelitis (CREAE) mice, a model of multiple sclerosis (MS). O-2093, either synthesized by us or commercially available, was inactive in the 'tetrad' up to a 20 mg kg(-1) dose (i.v.). Like O-2093, the other four compounds exhibited low affinity in CB(1) (K(i) from 1.3 to >10 microM) and CB(2) binding assays (1.310 microM), very low potency as fatty acid amide hydrolase (FAAH) inhibitors (IC(50)>25 microM) and were inactive in the 'tetrad' up to a 30 mg kg(-1) dose (i.v.). While O-2247 and O-2248 were poor inhibitors of [(14)C]AEA cellular uptake (IC(50)>40 microM), O-3246 and O-3262 were quite potent in this assay. O-3246, which exhibits only a very subtle structural difference with O-2093, is the most potent inhibitor of AEA uptake reported in vitro under our experimental conditions (IC(50)=1.4 microM) and is 12-fold more potent than O-2093. When injected intravenously O-3246 and O-3262, again like O-2093 and unlike O-2247 and O-2248, significantly inhibited limb spasticity in mice with CREAE. These data confirm the potential utility of selective AEA uptake inhibitors as anti-spasticity drugs in MS and, given the very subtle chemical differences between potent and weak inhibitors of uptake, support further the existence of a specific mechanism for this process.

Topics: Animals; Arachidonic Acids; Cell Line; Disease Models, Animal; Endocannabinoids; Mice; Multiple Sclerosis; Muscle Spasticity; Neuromuscular Blocking Agents; Polyunsaturated Alkamides; Rats

Endocannabinoids are released after brain injury and believed to attenuate neuronal damage by binding to CB(1) receptors and protecting against excitotoxicity. Such excitotoxic brain lesions initially result in primary destruction of brain parenchyma, which attracts macrophages and microglia. These inflammatory cells release toxic cytokines and free radicals, resulting in secondary neuronal damage. In this study, we show that the endocannabinoid system is highly activated during CNS inflammation and that the endocannabinoid anandamide (AEA) protects neurons from inflammatory damage by CB(1/2) receptor-mediated rapid induction of mitogen-activated protein kinase phosphatase-1 (MKP-1) in microglial cells associated with histone H3 phoshorylation of the mkp-1 gene sequence. As a result, AEA-induced rapid MKP-1 expression switches off MAPK signal transduction in microglial cells activated by stimulation of pattern recognition receptors. The release of AEA in injured CNS tissue might therefore represent a new mechanism of neuro-immune communication during CNS injury, which controls and limits immune response after primary CNS damage.

Topics: Adult; Animals; Arachidonic Acids; Cannabinoid Receptor Modulators; Cell Cycle Proteins; Cell Survival; Cells, Cultured; Dual Specificity Phosphatase 1; Encephalitis; Endocannabinoids; Extracellular Signal-Regulated MAP Kinases; Humans; Immediate-Early Proteins; MAP Kinase Signaling System; Mice; Microglia; Middle Aged; Multiple Sclerosis; Neurons; Neuroprotective Agents; Nitric Oxide; Nitric Oxide Synthase; Phosphoprotein Phosphatases; Phosphorylation; Polyunsaturated Alkamides; Protein Phosphatase 1; Protein Tyrosine Phosphatases; Rats; Rats, Wistar; Receptors, Cannabinoid; Tissue Distribution

Theiler's virus infection of the central nervous system (CNS) induces an immune-mediated demyelinating disease in susceptible mouse strains and serves as a relevant infection model for human multiple sclerosis (MS). Cannabinoids have been shown to exert beneficial effects on animal models of MS and evidence suggests that the endocannabinoid system plays a role in the tonic control of spasticity. In this study we show that OMDM1 [(R)-N-oleoyl-(1'-hydroxybenzyl)-2'-ethanolamine] and OMDM2 [(S)-N-oleoyl-(1'-hydroxybenzyl)-2'-ethanolamine], two selective inhibitors of the putative endocannabinoid transporter and hence of endocannabinoid inactivation, provide an effective therapy for Theiler murine encephalomyelitis virus-induced demyelinating disease (TMEV-IDD). Treatment of TMEV-infected mice with OMDM1 and OMDM2 enhanced anandamide levels in the spinal cord and ameliorated motor symptoms. This was associated with a down-regulation of inflammatory responses in the spinal cord. In addition we show that OMDM1 and OMDM2 down-regulate macrophage function by (i) decreasing the surface expression of major histocompatibility complex (MHC) class II molecules, (ii) inhibiting nitric oxide synthase-2 (NOS-2) expression and (iii) reducing the production of the pro-inflammatory cytokines interleukin-1beta (IL-1beta) and interleukin-12 (IL-12p40). Taken together, these results point to the manipulation of the endocannabinoid system as a possible strategy to develop future MS therapeutic drugs.

Topics: Animals; Arachidonic Acids; Benzyl Compounds; Cannabinoid Receptor Modulators; Cardiovirus Infections; Carrier Proteins; Cytokines; Disease Models, Animal; Endocannabinoids; Female; Histocompatibility Antigens Class II; Inflammation; Inflammation Mediators; Macrophages; Mice; Microglia; Motor Activity; Multiple Sclerosis; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Polyunsaturated Alkamides; Theilovirus; Up-Regulation

Recent studies have addressed the changes in endocannabinoid ligands and receptors that occur in multiple sclerosis, as a way to explain the efficacy of cannabinoid compounds to alleviate spasticity, pain, tremor, and other signs of this autoimmune disease. Using Lewis rats with experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, we recently found a decrease in cannabinoid CB1 receptors mainly circumscribed to the basal ganglia, which could be related to the motor disturbances characteristic of these rats. In the present study, using the same model, we explored the potential changes in several neurotransmitters in the basal ganglia that might be associated with the motor disturbances described in these rats, but we only found a small increase in glutamate contents in the globus pallidus. We also examined whether the motor disturbances and the changes of CB1 receptors found in the basal ganglia of EAE rats disappear after the treatment with rolipram, an inhibitor of type IV phosphodiesterase able to supress EAE in different species. Rolipram attenuated clinical decline, reduced motor inhibition, and normalized CB1 receptor gene expression in the basal ganglia. As a third objective, we examined whether EAE rats also exhibited changes in endocannabinoid levels as shown for CB1 receptors. Anandamide and 2-arachidonoylglycerol levels decreased in motor related regions (striatum, midbrain) but also in other brain regions, although the pattern of changes for each endocannabinoid was different. Finally, we hypothesized that the elevation of the endocannabinoid activity, following inhibition of endocannabinoid uptake, might be beneficial in EAE rats. AM404, arvanil, and OMDM2 were effective to reduce the magnitude of the neurological impairment in EAE rats, whereas VDM11 did not produce any effect. The beneficial effects of AM404 were reversed by blocking TRPV1 receptors with capsazepine, but not by blocking CB1 receptors with SR141716, thus indicating the involvement of endovanilloid mechanisms in these effects. However, a role for CB1 receptors is supported by additional data showing that CP55,940 delayed EAE progression. In summary, our data suggest that reduction of endocannabinoid signaling is associated with the development of EAE in rats. We have also proved that the reduction of CB1 receptors observed in these rats is corrected following treatment with a compound used in EAE such as rolipram. In addition, the direct or i

Topics: 3',5'-Cyclic-AMP Phosphodiesterases; Animals; Arachidonic Acids; Basal Ganglia; Brain; Cannabinoid Receptor Modulators; Capsaicin; Cyclic Nucleotide Phosphodiesterases, Type 4; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Endocannabinoids; Gene Expression; Glycerides; Male; Multiple Sclerosis; Phosphodiesterase Inhibitors; Polyunsaturated Alkamides; Rats; Rats, Inbred Lew; Receptor, Cannabinoid, CB1; Receptors, Cannabinoid; Rolipram; TRPV Cation Channels

Spasticity is a complicating sign in multiple sclerosis that also develops in a model of chronic relapsing experimental autoimmune encephalomyelitis (CREAE) in mice. In areas associated with nerve damage, increased levels of the endocannabinoids, anandamide (arachidonoylethanolamide, AEA) and 2-arachidonoyl glycerol (2-AG), and of the AEA congener, palmitoylethanolamide (PEA), were detected here, whereas comparable levels of these compounds were found in normal and non-spastic CREAE mice. While exogenously administered endocannabinoids and PEA ameliorate spasticity, selective inhibitors of endocannabinoid re-uptake and hydrolysis-probably through the enhancement of endogenous levels of AEA, and, possibly, 2-arachidonoyl glycerol-significantly ameliorated spasticity to an extent comparable with that observed previously with potent cannabinoid receptor agonists. These studies provide definitive evidence for the tonic control of spasticity by the endocannabinoid system and open new horizons to therapy of multiple sclerosis, and other neuromuscular diseases, based on agents modulating endocannabinoid levels and action, which exhibit little psychotropic activity.

Topics: Amides; Animals; Arachidonic Acids; Brain; Cannabinoid Receptor Modulators; Cannabinoids; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Endocannabinoids; Ethanolamines; Glycerides; Humans; Mice; Mice, Inbred Strains; Multiple Sclerosis; Palmitic Acids; Piperidines; Polyunsaturated Alkamides; Pyrazoles; Receptors, Cannabinoid; Receptors, Drug; Rimonabant; Spasm; Spinal Cord