Use of designer GPCRs to study GPCR regulation of key metabolic pathways Armbruster et al. (PNAS 104, 5163-8, 2007) first described a set of muscarinic receptor-based designer GPCRs which are now generally referred to as DREADDs ('designer receptors exclusively activated by designer drugs'). These designer receptors are unable to bind the endogenous muscarinic receptor agonist, acetylcholine, due to two single point mutations introduced into the transmembrane receptor core. Importantly, DREADDs can be efficiently activated by a compound called clozapine-N-oxide (CNO), an agent that is otherwise pharmacologically inert. The first DREADDs that were developed represent GPCRs that selectively activate G proteins of the Gq or Gi family, respectively. We recently generated additional DREADDs endowed with different coupling properties. For example, we designed an M3 muscarinic receptor (M3R)/beta1-adrenergic receptor hybrid DREADD that is able to selectively activate Gs (Guettier et al., PNAS 106, 19197-202, 2009). More recently, we also generated an M3R-based DREADD that is unable to couple to G proteins but retains arrestin-dependent signaling (Nakajima and Wess, Mol Pharmacol 82, 575-82, 2012). We are currently in the process of expressing DREADDs with different coupling properties in metabolically relevant cell types. These cell types include adipocytes, pancreatic beta-cells, skeletal muscle cells, and hepatocytes. We are also using flex switch technology to selectively express various DREADDs in distinct neuronal subpopulations of the hypothalamus. Preliminary results indicate that CNO treatment of some of these mutant mouse strains has pronounced effect on glucose and energy homeostasis (K. Nakajima, M. Rossi, L. Zhu, D. Bone, Z. Cui, et al.; unpublished results). The following paragraph summarizes a recent study in which we used DREADD technology to explore the role of Gq signaling in regulating hepatic glucose production. Activation of Gq signaling in hepatocytes promotes hepatic glucose production Elevated hepatic glucose production (HGP) is the major contributor to fasting hyperglycemia in T2D. For this reason, a better understanding of the signaling pathways that regulate hepatic glucose fluxes is of great potential clinical relevance. At present, the in vivo metabolic roles of hepatic Gq-coupled GPCRs remain poorly understood. To address this issue, we generated a transgenic mouse line that expresses the M3R-based Gq DREADD in selectively in hepatocytes (Hep-Rq mice). For the sake of simplicity, we refer to this Gq DREADD simply as 'Rq'. Acute treatment of Hep-Rq mice with CNO caused pronounced, dose-dependent increases in blood glucose levels. Moreover, CNO-injected Hep-Rq mice showed impaired glucose tolerance and significantly increased blood glucose levels in a pyruvate challenge test, consistent with an Rq-mediated stimulation of HGP. The Rq-mediated increase in HGP was similar in magnitude to that observed after treatment of Hep-Rq mice with glucagon which promotes HGP via Gs-dependent mechanisms. Isotope-labeling studies indicated that Gq signaling leads to enhanced HGP by stimulating both gluconeogenesis and glycogenolysis. These findings suggest that Gq-linked GPCRs play a critical role in regulating hepatic glucose fluxes. We also demonstrated that the expression levels of several Gq-coupled receptors were significantly increased in the liver of leptin-deficient mice (ob/ob mice), as compared to lean littermates. Among all GPCR genes analyzed, the V1b vasopressin receptor showed the most robust increase in receptor transcript levels. Strikingly, in vivo studies demonstrated that treatment of ob/ob mice with SSR149415, a selective V1b receptor antagonist, was able to significantly reduce the increase in HGP displayed by leptin-deficient mice. We are currently further exploring the possibility that V1b vasopressin antagonists might prove useful to ameliorate the metabolic deficits associated with obesity and T2D.