The G protein-coupled receptor 83 (Gpr83) is widely expressed in brain regions regulating energy metabolism. Here we report that hypothalamic expression of Gpr83 is regulated in response to nutrient availability and is decreased in obese mice compared with lean mice. In the arcuate nucleus, Gpr83 colocalizes with the ghrelin receptor (Ghsr1a) and the agouti- related protein. In vitro analyses show heterodimerization of Gpr83 with Ghsr1a diminishes activation of Ghsr1a by acyl-ghrelin. The orexigenic and adipogenic effect of ghrelin is accordingly potentiated in Gpr83-deficient mice. Interestingly, Gpr83 knock-out mice have normal body weight and glucose tolerance when fed a regular chow diet, but are protected from obesity and glucose intolerance when challenged with a high-fat diet, despite hyper- phagia and increased hypothalamic expression of agouti-related protein, Npy, Hcrt and Ghsr1a. Together, our data suggest that Gpr83 modulates ghrelin action but also indicate that Gpr83 regulates systemic metabolism through other ghrelin-independent pathways.
The murine G protein-coupled receptor (GPCR) 83 (Gpr83) is an orphan receptor belonging to the rhodopsin-like or testis, as well as the white and brown adipose tissue (Fig. 1a). Of appreciable note, analysis of hypothalamic Gpr83 expression revealed a decrease in mice with diet-induced obesity as compared with age-matched, lean controls (Fig. 1b). Further- more, fasting of lean mice for 12, 24 or 36 h resulted in a time- dependent decrease in the hypothalamic expression of Gpr83 (Fig. 1c). Refeeding with either a HFD or a fat-free diet for 6 h caused a subsequent increase in hypothalamic Gpr83 mRNA expression (Fig. 1c). In situ hybridization histochemistry in combination with immunohistochemistry revealed a broad abundance of Gpr83 in the ARC. In particular, we found Gpr83 expressed in specific subsets of ARC neurons that express Ghsr1a (Fig. 2a–c) and AgRP (Fig. 2d,f).
Heterodimerization of Gpr83 and Ghsr1a. To determine whether Gpr83 influences ghrelin–Ghsr1a signalling, we next examined whether both receptors interact in vitro. Both sandwich enzyme-linked immunosorbent assay (ELISA) and a yellow fluorescent protein (YFP)-based protein complementation assay (YFP-PCA) revealed heterodimerization of Gpr83 and Ghsr1a (Fig. 3a,b). Furthermore, cotransfection of Ghsr1a and Gpr83 in HEK293 cells led to a 43% reduction in ghrelin-stimulated Ghsr1a activity compared with control cells cotransfected with Ghsr1a and the rat muscarinic acetylcholine receptor M3 (rM3R) (Fig. 3c). Of appreciable note, this effect was not due to changes in the cell surface or total Ghsr1a expression (Fig. 3d,e), and Gpr83 activity was not influenced by treatment with acyl ghrelin, des-acyl ghrelin or obestatin (Supplementary Fig. S1A). Furthermore, heterodimerization of Gpr83 and Ghsr1a was also observed in mouse hypothalamic N41 cells cotransfected with Ghsr1a and Gpr83 (Supplementary Fig. S1B). Together, these data indicate that Gpr83–Ghsr1a heterodimerization reduces activation of Ghsr1a by ghrelin, thus supporting a possible role for Gpr83 in the regulation of energy homeostasis by reducing the ability of ghrelin to activate its only known endogenous receptor.
Metabolic phenotype of chow-fed Gpr83 ko mice. To determine whether Gpr83 has a role in regulating systemic metabolism in vivo, we next assessed the metabolic phenotype of Gpr83- deficient (Gpr83 ko) mice. These mice had normal body weight, food intake, glucose tolerance and insulin sensitivity when fed a regular chow diet (Fig. 4), whereas free-fatty acids (FFA) were increased under the same conditions (Table 1). Compared with wild-type (wt) control mice, body fat mass (Fig. 4b) and plasma leptin levels (Table 1) were moderately decreased without any1 family A of GPCRs . This family is by far the largest subfamily of GPCRs and includes several receptors implicated in the regulation of systemic metabolism, such as the ghrelin receptor (Ghsr1a)2,3 and the orexin receptors 1 and 2 (Ox1r and Ox2r)4. Gpr83 was originally identified as a glucocorticoid- induced transcript in a murine T cell line and, therefore, was referred to as glucocorticoid-induced receptor5. Induction of Gpr83 mRNA expression following dexamethasone6 or amphetamine7 treatment suggests a possible role in the regu- lation of the hypothalamus–pituitary–adrenal axis. This possibility is further supported by the observation that Gpr83 is expressed in hypothalamic nuclei that govern energy balance, such as the arcuate nucleus (ARC), the paraventricular nucleus and the lateral hypothalamic area6,8–10. As Gpr83 is expressed in hypothalamic nuclei relevant for energy metabolism control and as a potential modulator of the hypothalamus–pituitary–adrenal axis might also be relevant for systems metabolism, we hypothesized that Gpr83 might have a role in the central regulation of energy metabolism.
To test this hypothesis, we analysed the expression of Gpr83 in the mouse hypothalamus and observed that its expression is regulated by nutrient availability. In the ARC, we further find Gpr83 being colocalized with both the ghrelin receptor (Ghsr1a) and the agouti-related protein (AgRP), and in vitro analysis in HEK293 and mouse hypothalamic N41 cells show hetero- dimerization of Gpr83 with Ghsr1a that reduces ghrelin receptor activity. In line with this observation, the effects of ghrelin treatment on food intake and adiposity are enhanced in Gpr83 knock-out (Gpr83 ko) mice, thus supporting a functional interaction between Gpr83 and Ghsr1a in vivo. However, Gpr83 ko mice are also protected from obesity and glucose intolerance following exposure to high-fat diet (HFD), despite relative hyperphagia and increased hypothalamic expression of AgRP, Npy, Hcrt and Ghsr1a. Taken together, these data suggest that Gpr83 has multiple ghrelin-dependent and ghrelin-independent roles in the regulation of systemic energy metabolism.