Enhancing endocannabinoid tone reduces neural stem cell proliferation: a role for interleukin-1β

An emerging concept in the field of neural stem cell biology is that endocannabinoids (eCB) in the central nervous system (CNS) play a modulatory role in the development of the CNS as they are able to attenuate certain detrimental effects of neuroinflammation on neurogenesis throughout adult life. The dynamic interplay between the endocannabinoid system and the CNS offers great potential insights into the neurochemical underpinnings of neural degeneration. Neural stem cells (NSC) have the possibility of replacing lost CNS cells during neurogenesis. However, they are limited by pro-inflammatory cytokines during neurodegeneration. Interleukin-6 (IL-6) is a pleiotropic cytokine with both anti- and pro-inflammatory properties. It is involved in neurogenesis, neural regeneration and metabolic regulation. Interferon gamma-induced protein 10 (IP10) is a pro-inflammatory cytokine involved in the regulation of immune responses. Binding of IP10 to the receptor CXCR3 can lead to a multitude of physiological responses such as cell growth, chemotaxis and apoptosis. Tumour Necrosis Factor (TNF-ɑ) is a monocyte-derived cytotoxin involved in systemic inflammation and in the regulation of NSC migration, proliferation and self-renewal. Growing evidence suggests that TNF-ɑ signalling pathways interact with the endocannabinoid system in a bidirectional fashion to control neural plasticity and repair.

Aims. The present study examined the effects of inhibiting the primary enzymes responsible for the metabolism of the endocannabinoids 2-AG and anandamide, namely monoacylglycerol lipase (MAGL) and fatty acid amide hydrolase (FAAH) respectively, on neural stem cell proliferation, NSC viability, and cytokine release.

Methods. NSC cultures prepared from the cortex of embryonic day 16 (ED16) C57BL6 mice (wild type, WT) and IL-1β knock-out (KO) mice. NSC were treated with FAAH inhibitor URB597 (1.5 µM) or MAGL inhibitor JZL184 (3 µM) and incubated for 24h. MTT and LDH labelling was assessed to determine the effect of exposure of NSCs to URB-597 and JZL-184 on cell proliferation or cell death. Finally, cytokine release was assessed using sandwich ELISA kits for IP10-, IL6- and TNF-α, both in wildtype and IL-1β KO NSC cultures.

Results. Exposure of WT NSCs to URB-597 (n=8) or JZL-184 (n=8) induced a significant reduction in proliferation when compared to the control group (p<0.01). In comparison, an increase in proliferation was observed in IL1-β KO NSCs treated with URB597 (n=8) or JZL184 (n=8) (p<0.01). URB-597 or JZL-184 did not significant alter cell viability in either WT or IL1-β KO NSCs. WT NSCs exposed to URB-597 showed a significant increase in the release of TNF (n=8, p<0.0001), IP10 (n=8, p=0.0433) and IL-6 (n=8, p=0.0311). Interestingly, this effect was not observed in IL1-β KO NSCs. Exposure of WT NSCs to JZL184 resulted in increases in IP10 release (n=8, p=0.0307), an effect enhanced in IL1-β KO NSCs (n=8, p=0.0021).

Conclusions. The findings of this study demonstrate that enhancing endocannabinoid tone by inhibiting the enzymes which degradate anandamide and 2-AG enhances pro-inflamamtory cytokine release and decreases NSC proliferation. In comparison, in the absence of IL1-β signalling, NSC proliferation is enhanced following identical treatment regimes, suggesting an important role for IL-1β in mediating the effect of endocannabinoids on neurogenesis.

URB-597 (NIMH C-919) and JZL-184 (NIMH N-906) were kindly provided by the NIMH Chemical Synthesis and Drug Supply Program (USA). Supported by The Royal Society, UK (International Exchange Programme ref: IE131551e)