JWH-015

We tested the efficacy of 4'-fluorocannabidiol (4'-F-CBD), a semisynthetic cannabidiol derivative, and HU-910, a cannabinoid receptor 2 (CB2) agonist in resolving l-DOPA-induced dyskinesia (LID). Specifically, we were interested in studying whether these compounds could restrain striatal inflammatory responses and rescue glutamatergic disturbances characteristic of the dyskinetic state. C57BL/6 mice were rendered hemiparkinsonian by unilateral striatal lesioning with 6-OHDA. Abnormal involuntary movements were then induced by repeated i.p. injections of l-DOPA + benserazide. After LID was installed, the effects of a 3-day treatment with 4'-F-CBD or HU-910 in combination or not with the TRPV1 antagonist capsazepine (CPZ) or CB2 agonists HU-308 and JWH015 were assessed. Immunostaining was conducted to investigate the impacts of 4'-F-CBD and HU-910 (with CPZ) on inflammation and glutamatergic synapses. Our results showed that the combination of 4'-F-CBD + CPZ, but not when administered alone, decreased LID. Neither HU-910 alone nor HU-910+CPZ were effective. The CB2 agonists HU-308 and JWH015 were also ineffective in decreasing LID. Both combination treatments efficiently reduced microglial and astrocyte activation in the dorsal striatum of dyskinetic mice. However, only 4'-F-CBD + CPZ normalized the density of glutamate vesicular transporter-1 (vGluT1) puncta colocalized with the postsynaptic density marker PSD95. These findings suggest that 4'-F-CBD + CPZ normalizes dysregulated cortico-striatal glutamatergic inputs, which could be involved in their anti-dyskinetic effects. Although it is not possible to rule out the involvement of anti-inflammatory mechanisms, the decrease in striatal neuroinflammation markers by 4'-F-CBD and HU-910 without an associated reduction in LID indicates that they are insufficient per se to prevent LID manifestations.

Neuroinflammation is closely related to the pathogenesis of neurodegenerative diseases. Activation of microglia, the resident immune cells in CNS, induces inflammatory responses, resulting in the release of neurotoxic molecules, which favors neuronal death and neurodegeneration. Nuclear receptor-related 1 (Nurr1) protein, one of the orphan nuclear receptor superfamilies, is an emerging target for neuroprotective therapy. In addition, the anti-inflammatory function of cannabinoid (CB) receptors has attracted increasing interest. As both CB receptors (especially CB2 receptor) and Nurr1 exist in microglia, and regulate a number of same molecular points such as NF-κB, we herein explored the interplay between the CB2 receptor and Nurr1 as well as the regulatory mechanisms in microglial cells. We showed that the application of CB2 receptor agonists JWH015 (1, 10 μM) significantly increased the nuclear Nurr1 protein in BV-2 cells and primary midbrain microglia. Overexpression of Nurr1 or application of Nurr1 agonist C-DIM12 (10 μM) significantly increased the mRNA level of CB2 receptor in BV-2 cells, suggesting that positive expression feedback existing between the CB2 receptor and Nurr1. After 2-AG and JWH015 activated the CB2 receptors, the levels of p-ERK, p-AKT, p-GSK-3β in BV-2 cells were significantly increased. Using ERK1/2 inhibitor U0126 and PI3K/AKT inhibitor LY294002, we revealed that the amount of Nurr1 in the nucleus was upregulated through β-arrestin2/ERK1/2 and PI3K/AKT/GSK-3β signaling pathways. With these inhibitors, we found a cross-talk interaction between the two pathways, and the ERK1/2 signaling pathway played a more dominant regulatory role. Furthermore, we demonstrated that when the CB2 receptor was activated, the phagocytic function of BV-2 cells was significantly weakened; the activation of Nurr1 also inhibited the phagocytic function of BV-2 cells. Pretreatment with the signaling pathway inhibitors, especially U0126, reversed the inhibitory effect of 2-AG on phagocytosis, suggesting that CB2 receptor may regulate the phagocytic function of microglia by activating Nurr1. In conclusion, CB2 receptor or/and Nurr1-mediated signal pathways play instrumental roles in the progress of phagocytosis, which are expected to open up new treatment strategies for neurodegenerative diseases.