Study of the effects induced by GH receptor ablation in neurons of the lateral hypothalamic area

Abstract

The Lateral Hypothalamic Area (LHA) contains several neural populations that control important vegetative and metabolic functions, including food intake, energy expenditure, autonomic control, and several cognitive aspects, such as wakefulness-sleep cycle and reward system. Therefore, the understanding of how hormonal factors affect the functioning and activity of these neurons is fundamental in the study of neuroendocrine and metabolic disorders. Our research group demonstrated that a large number of LHA neurons are directly responsive to Growth Hormone (GH). In addition, recent works have shown that the action of GH in the central nervous system has an important effect on metabolic and cognitive aspects. However, no study has investigated the importance of GH action specifically on LHA so far. Thus, the objective of this postdoctoral project is to investigate the effects induced by the ablation of the GH receptor (GHR) on LHA neurons. For this, stereotactic injection of viral vector (AAV) which induces expression of the Cre enzyme in mice presenting the Ghr gene flanked by LoxP sites will be used. This way, the GHR will be inactivated only in the area that will be infected by the virus. The injections will be performed bilaterally on male mice of 8 to 10 weeks old and will be centered on LHA. Control animals will receive a viral vector injection that induces only GFP expression. Preliminary experiments will help to determine the exact coordinates of the injection, taking as a starting point values obtained in mice neuroanatomical atlases. After one week of recovery and for the induction of gene recombination, the animals will be followed weekly to detect possible changes in weight and body composition. Several metabolic variables will be measured including analyses of food intake, energy expenditure, ambulatory activity, respiratory quotient, glucose tolerance and insulin sensitivity. The experimental animals will also be evaluated regarding their sensitivity to the action of hormones that act through LHA, such as leptin. Additional experiments with animals receiving a high-fat diet or undergoing food deprivation for metabolic analyzes will be conducted on additional groups of animals. In addition, the wakefulness-sleep pattern and palatability to pleasant foods (hyperlipid diet and sucrose solution) will be evaluated. Additional behavioral analyses may be conducted according to the initial results. At the end of the experiments, the injection site in all animals will be histologically inspected and confirmation of gene deletion will occur due to the absence of a GH injection capacity to induce STAT5 phosphorylation, a marker that has been used to indicate functional expression of GHR. In summary, the results of the proposed experiments will provide original and potentially relevant data that can improve our understanding of the central action of GH. Thus, this study will contribute to a better understanding of endocrine and metabolic problems caused by changes in GH secretion and the participation of LHA in these effects. (AU)