Synucleinopathies such as Parkinson's disease are characterized by the pathological deposition of misfolded a-synuclein aggregates into inclusions throughout the central and peripheral nervous system. Mounting evidence suggests that intercellular propagation of a-synuclein aggregates may contribute to the neuropathology; however, the mechanism by which spread occurs is not fully understood. By using quantitative fluorescence microscopy with co-cultured neurons, here we show that a-synuclein fibrils efficiently transfer from donor to acceptor cells through tunneling nanotubes (TNTs) inside lysosomal vesicles. Following transfer through TNTs, a-synuclein fibrils are able to seed soluble a-synuclein aggregation in the cytosol of acceptor cells. We propose that donor cells overloaded with a-synuclein aggregates in lysosomes dispose of this material by hijacking TNT-mediated intercellular trafficking. Our findings thus reveal a possible novel role of TNTs and lysosomes in the progression of synucleinopathies.

Recent evidence suggests that disease progression in Parkinson's disease (PD) could occur by the spreading of α-synuclein (α-syn) aggregates between neurons. Here we studied the role of astrocytes in the intercellular transfer and fate of α-syn fibrils, using in vitro and ex vivo models. α-Syn fibrils can be transferred to neighboring cells; however, the transfer efficiency changes depending on the cell types. We found that α-syn is efficiently transferred from astrocytes to astrocytes and from neurons to astrocytes, but less efficiently from astrocytes to neurons. Interestingly, α-syn puncta are mainly found inside the lysosomal compartments of the recipient cells. However, differently from neurons, astrocytes are able to efficiently degrade fibrillar α-syn, suggesting an active role for these cells in clearing α-syn deposits. Astrocytes co-cultured with organotypic brain slices are able to take up α-syn fibrils from the slices. Altogether our data support a role for astrocytes in trapping and clearing α-syn pathological deposits in PD.

The endocannabinoid system exhibits anti-inflammatory properties by regulating cytokine production. Anandamide (AEA) down-regulates proinflammatory cytokines in a viral model of multiple sclerosis (MS). However, little is known about the mechanisms by which AEA exerts these effects. Microglial cells are the main source of cytokines within the brain and the first barrier of defense against pathogens by acting as antigen presenting cells. IL-10 is a key physiological negative regulator of microglial activation. In this study we show that AEA enhances LPS/IFNgamma-induced IL-10 production in microglia by targeting CB(2) receptors through the activation of ERK1/2 and JNK MAPKs. AEA also inhibits NF-kappaB activation by interfering with the phosphorylation of IkappaBalpha, which may result in an increase of IL-10 production. Moreover, endogenously produced IL-10 negatively regulates IL-12 and IL-23 cytokines, which in its turn modify the pattern of expression of transcription factors involved in Th commitment of splenocytes. This suggests that by altering the cytokine network, AEA could indirectly modify the type of immune responses within the central nervous system (CNS). Accordingly, pharmacological modulation of AEA uptake and degradation might be a useful tool for treating neuroinflammatory diseases.

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The endocannabinoid anandamide (AEA) is released by macrophages and microglia on pathological neuroinflammatory conditions such as multiple sclerosis (MS). CD200 is a membrane glycoprotein expressed in neurons that suppresses immune activity via its receptor (CD200R) mainly located in macrophages/microglia. CD200‐CD200R interactions contribute to the brain immune privileged status. In this study, we show that AEA protects neurons from microglia‐induced neurotoxicity via CD200‐CD200R interaction. AEA increases the expression of CD200R1 in LPS/IFN‐γ activated microglia through the activation of CB2 receptors. The neuroprotective effect of AEA disappears when microglial cells derive from CD200R1−/− mice. We also show that engagement of CD200R1 by CD200Fc decreased the production of the proinflammatory cytokines IL‐1β and IL‐6, but increased IL‐10 in activated microglia. In the chronic phases of Theiler's virus‐induced demyelinating disease (TMEV‐IDD) the expression of CD200 and CD200R1 was reduced in the spinal cord. AEA‐treated animals up‐regulated the expression of CD200 and CD200R1, restoring levels found in sham animals together with increased expression of IL‐10 and reduced expression of IL‐1β and IL‐6. Treated animals also improved their motor behavior. Because AEA up‐regulated the expression of CD200R1 in microglia, but failed to enhance CD200 in neurons we suggest that AEA‐induced up‐regulation of CD200 in TMEV‐IDD is likely due to IL‐10 as this cytokine increases CD200 in neurons. Our findings provide a new mechanism of action of AEA to limit immune response in the inflamed brain. © 2012 Wiley Periodicals, Inc.

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Cannabinoids have recently been approved as a treatment for pain in multiple sclerosis (MS). Increasing evidence from animal studies suggests that this class of compounds could also prove efficient to fight neurodegeneration, demyelination, inflammation and autoimmune processes occurring in this pathology. However, the use of cannabinoids is limited by their psychoactive effects. In this context, potentiation of the endogenous cannabinoid signalling could represent a substitute to the use of exogenously administrated cannabinoid ligands. Here, we studied the expression of different elements of the endocannabinoid system in a chronic model of MS in mice. We first studied the expression of the two cannabinoid receptors, CB1 and CB2, as well as the putative intracellular cannabinoid receptor peroxisome proliferator‐activated receptor‐α. We observed an upregulation of CB2, correlated to the production of proinflammatory cytokines, at 60 days after the onset of the MS model. At this time, the levels of the endocannabinoid, 2‐arachidonoylglycerol, and of the anti‐inflammatory anandamide congener, palmithoylethanolamide, were enhanced, without changes in the levels of anandamide. These changes were not due to differences in the expression of the degradation enzymes, fatty acid amide hydrolase and monoacylglycerol lipase, or of biosynthetic enzymes, diacylglycerol lipase‐α and N‐acylphosphatidylethanolamine phospholipase‐D at this time (60 days). Finally, the exogenous administration of palmitoylethanolamide resulted in a reduction of motor disability in the animals subjected to this model of MS, accompanied by an anti‐inflammatory effect. This study overall highlights the potential therapeutic effects of endocannabinoids in MS.

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