Role of alpha7nAChR cholinergic receptor in the hypothalamic control of energy homeostasis in mice

Author(s)

Camilla Mendes de Souza

• Document type: Master's Dissertation
• Press: Limeira, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Faculdade de Ciências Aplicadas
• Defense date: 2019-08-07

Examining board members

Marcio Alberto Torsoni; Patrícia de Oliveira Prada; Daniela Soares Razolli

• Advisor: Marcio Alberto Torsoni

Abstract

Introduction: cholinergic signaling mediated by the activation of muscarinic and nicotinic receptors has been described in the literature as a classic and important signaling in the regulation of the inflammatory response. Recent research has investigated the role of acetylcholine, the physiological agonist of these receptors, in the control of energy homeostasis, at the central level. Studies have shown that mice that do not express acetylcholine in brain regions regulating energy homeostasis present excessive weight gain and hyperphagia. However, it has not yet been well described in the literature which cholinergic receptor subunits are involved in this response, making obscuration the signaling responsible for the observed effects. The hypothalamus is the regulating center of energy homeostasis and the 'alfa'7 subunit of the nicotinic acetylcholine receptor ('alfa'7nAChR) is highly expressed in this region. 'alfa'7nAChR when active recruits proteins such as JAK2 / STAT3 for its signaling, the same intracellular components required by leptin, anorexigenic hormone. The objective of the present study was to evaluate the role of the hypothalamic 'alfa'7nAChR receptor in the control of energy homeostasis. Methods: The work was developed with Swiss mice, under the authorization of CEUA / UNICAMP, registered under number 4787-1 / 2018. In brain sections, using the immunofluorescence technique, the presence of 'alfa'7nAChR receptors in hypothalamic cells regulating energy homeostasis was evaluated. The animals were submitted to stereotaxia in the lateral ventricle and intracerebroventricular stimulation (ICV) was used for administration of agonist (PNU) and antagonist ('alfa'bungarotoxin) of the receptor 'alfa'7nAChR and metabolic parameters were evaluated and in the hypothalamic tissue was evaluated the expression of neuropeptides by Real Time PCR and Western blot analysis. ICV-treated camogongs with siRNA, or inhibitors of JAK2 / STAT3 (AG490 and STATTIC) proteins, the expression of hypothalamic neuropeptides was evaluated. In Knockout animals for the 'alfa'7nAChR receptor, we evaluated food intake. Additionally in hypothalamic cell culture of mice, mypHo POMC-GFP lineage, we evaluated the expression of neuropeptides and pSTAT3 after stimulation with PNU. Results: Our immunofluorescence results indicate co-localization of 'alfa'7nAChR with 'alfa'-MSH, AgRP and NPY hypothalamic cells. Pharmacological activation of 'alfa'7nAChR reduced food intake and increased hypothalamic POMC expression and decreased NPY mRNA, AgRP and protein content of pAMPK. Inhibition of the 'alfa'7nAChR receptor with antagonist increased the mRNA content of NPY and AgRP. Inhibition of 'alfa'7nAChR with siRNA culminated in suppression of POMC expression and increase in AgRP mRNA levels. The knockout animals for 'alfa'7nAChR-/- showed no changes in food intake. Inhibition of proteins from the JAK2 / STAT3 signaling pathway reversed the effects observed under PNU stimulus. POMC-GFP cells, when treated with PNU, showed increased POMC expression and nuclear translocation of pSTAT3. Conclusion: selective activation of the 'alfa'7nAChR receptor is able to modulate important markers of response to food intake, suggesting that 'alfa'7nAChR activation can suppress the expression of orexigenic markers and favor the expression of anorectics using the intracellular JAK2 / STAT3 machinery (AU)

• FAPESP's process: 18/01863-6 - Role of hypothalamic cholinergic receptor alpha7 (alpha7nAChR) in control of energy homeostasis and association with leptin signaling in mice
• Grantee: Camilla Mendes de Souza
• Support Opportunities: Scholarships in Brazil - Master