Hypothalamic Pomc expression restricted to GABAergic neurons suppresses Npy overexpression and restores food intake in obese mice.

OBJECTIVE: Hypothalamic arcuate proopiomelanocortin (Arc-POMC) neurons are involved in different physiological processes such as the regulation of energy balance, glucose homeostasis, and stress-induced analgesia. Since these neurons heterogeneously express different biological markers and project to many hypothalamic and extrahypothalamic areas, it is proposed that Arc-POMC neurons could be classified into different subpopulations having diverse physiological roles. The aim of the present study was to characterize the contribution of the subpopulation of Arc-POMC neurons cosecreting gamma-aminobutyric acid (GABA) neurotransmitter in the control of energy balance. METHODS: Arc-Pomc expression restricted to GABAergic-POMC neurons was achieved by crossing a reversible Pomc-deficient mouse line (arcPomc-) with a tamoxifen-inducible Gad2-CreER transgenic line. Pomc expression was rescued in the compound arcPomc-/-:Gad2-CreER female and male mice by tamoxifen treatment at postnatal days 25 (P25) or 60 (P60), and body weight, daily food intake, fasting glycemia, and fasting-induced hyperphagia were measured. POMC recovery was quantified by immunohistochemistry and semiquantitative RT-PCR. Neuropeptide Y (NPY) and GABAergic neurons were identified by in situ hybridization. Arc-POMC neurons projecting to the dorsomedial hypothalamic nucleus (DMH) were studied by stereotactic intracerebral injection of fluorescent retrobeads into the DMH. RESULTS: Tamoxifen treatment of arcPomc-/-:Gad2-CreER mice at P60 resulted in Pomc expression in ∼23-25% of Arc-POMC neurons and ∼15-23% of Pomc mRNA levels, compared to Gad2-CreER control mice. Pomc rescue in GABAergic-POMC neurons at P60 normalized food intake, glycemia, and fasting-induced hyperphagia, while significantly reducing body weight. Energy balance was also improved in arcPomc-/-:Gad2-CreER mice treated with tamoxifen at P25. Distribution analysis of rescued POMC immunoreactive fibers revealed that the DMH is a major target site of GABAergic-POMC neurons. Further, the expression of the orexigenic neuropeptide Y (NPY) in the DMH was increased in arcPomc-/- obese mice but was completely restored after Pomc rescue in arcPomc-/-:Gad2-CreER mice. Finally, we found that ∼75% of Arc-POMC neurons projecting to the DMH are GABAergic. CONCLUSIONS: In the present study, we show that the expression of Pomc in the subpopulation of Arc-GABAergic-POMC neurons is sufficient to maintain normal food intake. In addition, we found that DMH-NPY expression is negatively correlated with Pomc expression in GABAergic-POMC neurons, suggesting that food intake may be regulated by an Arc-GABAergic-POMC → DMH-NPY pathway.

Hypothalamic Proopiomelanocortin Is Necessary for Normal Glucose Homeostasis in Female Mice.

The arcuate nucleus of the hypothalamus is a key regulator of energy balance and glucose homeostasis. In particular, arcuate proopiomelanocortin (POMC) neurons inhibit food intake, stimulate energy expenditure and increase glucose tolerance. The interruption of insulin or glucose signaling in POMC neurons leads to glucose intolerance without changing energy homeostasis. Although it was previously shown that POMC neurons are necessary for normal glucose homeostasis, the participation of POMC neuropeptide, by mechanisms independent of energy balance, remains to be demonstrated. To study the role of POMC in the regulation of glucose homeostasis, we performed glucose and insulin tolerance tests in non-obese mice lacking hypothalamic POMC expression. We found that POMC deficiency leads to glucose intolerance and insulin resistance in female mice before the onset of obesity or hyperphagia. Conversely, POMC deficiency does not impair glucose homeostasis in non-obese male mice. Interestingly, females completely normalize both glucose and insulin tolerance after genetic POMC restoration. Next, to further study sex dimorphism of POMC neurons regarding glucose homeostasis, we measured glucose-elicited changes in C-FOS by performing immunofluorescence in brain slices of POMC-EGFP mice. Remarkably, we found that glucose-induced C-FOS expression in POMC neurons is more than 3-fold higher in female than in male mice. Altogether, our results reveal a key role of arcuate POMC in the regulation of glucose homeostasis in females. Since POMC reactivation completely reverses the diabetogenic phenotype, arcuate POMC could be a potential target for diabetes therapy.