Role of alpha7 nicotinic acetylcholine receptor (alpha7nAchR) in preventing insulin resistance in hypothalamic cells

Neuronal hypothalamic insulin resistance is implicated with energy balance dysregulation and contributes to the pathogenesis of several neurodegenerative diseases. Its development has been intimately associated with a neuroinflammatory process mainly orchestrated by activated microglial cells. In this regard, our study aims to investigate a target highly expressed in the hypothalamus and involved in the regulation of the inflammatory process, but still poorly investigated within the context of insulin resistance: the ?7 nicotinic acetylcholine receptor (?7nAchR). To reproduce the physiological interactions that occur between neurons and microglia cells during the neuroinflammatory process in an in vitro model, the hypothalamic neuronal cell line mHypoA-2/29 was exposed to the conditioned medium obtained from BV-2 microglia cells treated with LPS. mHypoA-2/29 neurons exposed to the microglial conditioned medium (MCM) showed higher expression of the pro-inflammatory cytokines IL-6, IL-1? and TNF-?, in addition to developing insulin resistance. Activation of the ?7nAchR receptor with the selective agonist PNU-282987 prevented the inflammation induced by the conditioned medium, inhibiting NF-?B nuclear translocation and promoting an increase in IL-10 and tristetraprolin (TTP) gene expression. To confirm that these effects were indeed mediated by the ?7nAchR receptor, an mHypoA-2/29 knockout cell line for the gene encoding the ?7nAchR receptor (Chrna7) was developed via CRISPR/Cas9. In knockout cells, the effect of PNU to prevent the increase in the expression of pro-inflammatory cytokines and to promote the expression of IL-10 and TTP were lost. The anti-inflammatory action of the ?7nAchR receptor was also accompanied by an improvement in insulin sensitivity and a lower activation of neurodegeneration-related markers, such as GSK3 and tau. These effects were lost by the knockdown of Chrna7 via siRNA and by the pharmacological inhibition of the receptor and downstream proteins such as JAK2 and STAT3. Finally, we report that the deletion of the ?7nAchR receptor in mHypoA-2/29 neurons was marked by a pronounced reduction in cell viability and proliferation. Taken together, these results show that activation of ?7nAchR anti-inflammatory signaling in hypothalamic neurons exerts neuroprotective effects and prevents the development of insulin resistance induced by pro-inflammatory mediators secreted by microglial cells (AU)