G Protein-coupled Research Articles

Structural basis for the interaction between the Drosophila RTK Sevenless (dROS1) and the GPCR BOSS

Sevenless, the Drosophila homologue of ROS1 (University of Rochester Sarcoma) (herein, dROS1) is a receptor tyrosine kinase (RTK) essential for the differentiation of Drosophila R7 photoreceptor cells. Activation of dROS1 is mediated by binding to the extracellular region (ECR) of the GPCR (G protein coupled receptor) BOSS (Bride Of Sevenless) on adjacent cells. Activation of dROS1 by BOSS leads to subsequent downstream signaling pathways including SOS (Son of Sevenless). However, the physical basis for how dROS1 interacts with BOSS has long remained unknown. Here we provide a cryo-EM structure of dROS1’s extracellular region, which mediates ligand binding. We show that the extracellular region of dROS1 adopts a folded-over conformation stabilized by an N-terminal domain comprised of two disulfide stapled helical hairpins. We further narrowed down the interacting binding epitopes on both dROS1 and BOSS using hydrogen-deuterium exchange mass spectrometry (HDX-MS). This includes beta-strands in dROS1’s third Fibronectin type III (FNIII) domain and a C-terminal peptide in BOSS’ ECR. Our mutagenesis studies, coupled with AlphaFold complex predictions, support a binding interaction mediated by a hydrophobic interaction and beta-strand augmentation between these regions. Our findings provide a fundamental understanding of the regulatory function of dROS1 and further provide mechanistic insight into the human ortholog and oncogene ROS1.

Molecular architecture of human LYCHOS involved in lysosomal cholesterol signaling.

Lysosomal membrane protein LYCHOS (lysosomal cholesterol signaling) translates cholesterol abundance to mammalian target of rapamycin activation. Here we report the 2.11-Å structure of human LYCHOS, revealing a unique fusion architecture comprising a G-protein-coupled receptor (GPCR)-like domain and a transporter domain that mediates homodimer assembly. The NhaA-fold transporter harbors a previously uncharacterized intramembrane Na+ pocket. The GPCR-like domain is stabilized, by analogy to canonical GPCRs, in an inactive state through 'tethered antagonism' by a lumenal loop and strong interactions at the cytosol side preventing the hallmark swing of the sixth transmembrane helix seen in active GPCRs. A cholesterol molecule and an associated docosahexaenoic acid (DHA)-phospholipid are entrapped between the transporter and GPCR-like domains, with the DHA-phospholipid occupying a pocket previously implicated in cholesterol sensing, indicating inter-domain coupling via dynamic lipid-protein interactions. Our work provides a high-resolution framework for functional investigations of the understudied LYCHOS protein.

Development of a yeast-based sensor platform for evaluation of ligands recognized by the human Free Fatty Acid 2 Receptor.

Yeast-based sensors have shown great applicability for deorphanization of G protein-coupled receptors (GPCRs) and screening of ligands targeting these. A GPCR of great interest is free fatty acid 2 receptor (FFA2R), for which short-chain fatty acids such as propionate and acetate are agonists. FFA2R regulates a wide array of downstream receptor signaling pathways in both adipose tissue and immune cells and has been recognized as a promising therapeutic target, having been implicated in several metabolic and inflammatory diseases. While research aiming to identify ligands recognized by FFA2R for translational applications is ongoing, screening is complicated by the complex regulatory and cell-specific responses mediated by the receptor. To simplify screening towards identification of novel ligands, heterologous platforms are valuable tools that offer efficient identification of ligand activity in the absence of regulatory mechanisms. Here, we present a yeast-based sensor designed to evaluate G protein α i1 mediated FFA2R signaling, with an assay time of 3 h. We verify this platform towards the natural agonists, propionate and acetate, and show applicability towards evaluation of synthetic agonists, antagonists, and allosteric agonists. As such, we believe that the developed yeast strain constitutes a promising screening platform for effective evaluation of ligands acting on FFA2R.

Determination of the Negative Allosteric Binding Site of Cannabidiol at the CB1 Receptor: A Combined Computational and Site-Directed Mutagenesis Study.

Cannabinoid receptor 1 (CB1R) has been extensively studied as a potential therapeutic target for various conditions, including pain management, obesity, emesis, and metabolic syndrome. Unlike orthosteric agonists such as Δ9-tetrahydrocannabinol (THC), cannabidiol (CBD) has been identified as a negative allosteric modulator (NAM) of CB1R, among its other pharmacological targets. Previous computational and structural studies have proposed various binding sites for CB1R NAMs. An X-ray crystal structure revealed a binding site for the NAM, ORG27569, at an extrahelical location within the inner leaflet of the membrane. In contrast, multiple computational studies have previously proposed several potential allosteric binding sites for CBD within the CB1R structure. Given that a prior structural study suggested CBD might occupy the same site as ORG27569, we conducted a comprehensive investigation of potential CBD binding sites using molecular docking, molecular dynamics (MD) simulations, metadynamics (MTD) simulations, binding free-energy calculations, and in vitro mutagenesis experiments. Molecular docking, MD, and MTD simulations results, along with binding free-energy calculations, suggest that CBD may potentially bind to either the same extrahelical site as ORG27569 or a previously unidentified intracellular site located near TMHs 2, 6, and 7 and helix 8. This intracellular site is consistent with allosteric binding sites observed in other G protein-coupled receptors (GPCRs). To establish the most favorable allosteric site for CBD, we conducted site-directed mutagenesis of key residues at each site. Mutations at S4018.47ΔA and D4038.49ΔA augmented the binding of [3H]-SR141716A, suggesting these residues play critical roles in CBD binding. As a result, the combined computational and mutagenesis results identified a binding site for CBD between TMHs 2, 6, and 7 and helix 8, involving residues Y1532.40, I1562.43, M3376.29, L3416.33, S4018.47, and D4038.49. These findings provide valuable insights into how CBD binds to CB1R, thereby informing the rational design of new, selective, and potent NAMs. Moreover, the elucidation of this previously unexplored allosteric site might explain the polypharmacology of CBD due to structural conservation among Class A GPCRs.

FRPR-1, a G protein-coupled receptor (GPCR) in the FMRFamide-related peptide receptor family, modulates larval development as a receptor candidate of the FMRFamide-like peptide FLP-1 in Caenorhabditis elegans.

FMRFamide-like peptides (FLPs) and their receptors FMRFamide-related peptide receptors (FRPRs) are widely conserved in free-living and parasitic nematodes. Herein, we identified FRPR-1 as a of FLP-1 receptor candidate involved in larval development and diapause in the model nematode Caenorhabditis elegans. Our molecular genetic study, supported by in silico research, revealed the following: 1) frpr-1 loss-of-function completely suppresses the promotion of larval diapause caused by flp-1 overexpression; 2) AlphaFold2 analysis revealed the binding of FLP-1 to FRPR-1; 3) FRPR-1 as well as FLP-1modulates the production and secretion of the predominant insulin-like peptide DAF-28, which is produced in ASI neurons; and 4) the suppression of larval diapause by frpr-1 loss-of-function is completely suppressed by a daf-28 defect. Thus, FRPR-1 regulates larval development and diapause by modulating DAF-28 production and secretion. This study may provide new insights into the development of novel nematicides targeting parasitic nematodes using FRPR-1 inhibitors.

Systematical identification of regulatory GPCRs by single-cell trajectory inference reveals the role of ADGRD1 and GPR39 in adipogenesis.

Adipogenesis is the healthy expansion of white adipose tissue (WAT), serving as a compensatory response to maintain metabolic homeostasis in the presence of excess energy in the body. Therefore, the identification of novel regulatory molecules in adipogenesis, specifically membrane receptors such as G protein-coupled receptors (GPCRs), holds significant clinical promise. These receptors can serve as viable targets for pharmaceuticals, offering potential for restoring metabolic homeostasis in individuals with obesity. We utilized trajectory inference methods to analyze three distinct single-nucleus sequencing (sNuc-seq) datasets of adipose tissue and systematically identified GPCRs with the potential to regulate adipogenesis. Through verification in primary adipose progenitor cells (APCs) of mice, we discovered that ADGRD1 promoted the differentiation of APCs, while GPR39 inhibits this process. In the obese mouse model induced by a high-fat diet (HFD), both gain-of-function and loss-of-function studies validated that ADGRD1 promoted adipogenesis, thereby improving metabolic homeostasis, while GPR39 inhibited adipogenesis, leading to metabolic dysfunction. Additionally, through the analysis of 2,400 ChIP-seq data and 1,204 bulk RNA-seq data, we found that the transcription factors (TFs) MEF2D and TCF12 regulated the expression of ADGRD1 and GPR39, respectively. Our study revealed the regulatory role of GPCRs in adipogenesis, providing novel targets for clinical intervention of metabolic dysfunction in obese patients.

Conformational diversity in class C GPCR positive allosteric modulation

The metabotropic glutamate receptors (mGlus) are class C G protein-coupled receptors (GPCR) that form obligate dimers activated by the major excitatory neurotransmitter L-glutamate. The architecture of mGlu receptor comprises an extracellular Venus-Fly Trap domain (VFT) connected to the transmembrane domain (7TM) through a Cysteine-Rich Domain (CRD). The binding of L-glutamate in the VFTs and subsequent conformational change results in the signal being transmitted to the 7TM inducing G protein binding and activation. The mGlu receptors signal transduction can be allosterically potentiated by positive allosteric modulators (PAMs) binding to the 7TMs, which are of therapeutic interest in various neurological disorders. Here, we report the cryoEM structures of metabotropic glutamate receptor 5 (mGlu5) purified with three chemically and pharmacologically distinct PAMs. We find that the PAMs modulate the receptor equilibrium through their different binding modes, revealing how their interactions in the 7TMs impact the mGlu5 receptor conformational landscape and function. In addition, we identified a PAM-free but agonist-bound intermediate state that also reveals interactions mediated by intracellular loop 2. The activation of mGlu5 receptor is a multi-step process in which the binding of the PAMs in the 7TM modulates the equilibrium towards the active state.

Dysregulated autoantibodies targeting AGTR1 are associated with the accumulation of COVID-19 symptoms

Coronavirus disease 2019 (COVID-19) presents a wide spectrum of symptoms, the causes of which remain poorly understood. This study explored the associations between autoantibodies (AABs), particularly those targeting G protein-coupled receptors (GPCRs) and renin‒angiotensin system (RAS) molecules, and the clinical manifestations of COVID-19. Using a cross-sectional analysis of 244 individuals, we applied multivariate analysis of variance, principal component analysis, and multinomial regression to examine the relationships between AAB levels and key symptoms. Significant correlations were identified between specific AABs and symptoms such as fever, muscle aches, anosmia, and dysgeusia. Notably, anti-AGTR1 antibodies, which contribute to endothelial glycocalyx (eGC) degradation, a process reversed by losartan, have emerged as strong predictors of core symptoms. AAB levels increased with symptom accumulation, peaking in patients exhibiting all four key symptoms. These findings highlight the role of AABs, particularly anti-AGTR1 antibodies, in determining symptom severity and suggest their involvement in the pathophysiology of COVID-19, including vascular complications.

An Alternative Mode of GPCR Transactivation: Activation of GPCRs by Adhesion GPCRs

G protein-coupled receptors (GPCRs), critical for cellular communication and signaling, represent the largest cell surface protein family and play important roles in numerous pathophysiological processes. Consequently, GPCRs have become a primary focus in drug discovery efforts. Beyond their traditional G protein-dependent signaling pathways, GPCRs are also capable of activating alternative signaling mechanisms, including G protein-independent signaling, biased signaling, and signaling crosstalk. A particularly novel signaling mode employed by these receptors is GPCR transactivation, which enables cross-communication between GPCRs and other receptor types. Intriguingly, GPCR transactivation by distinct GPCRs has also been identified. In this review, I provide an overview of the known GPCR transactivation mechanisms and explore recently uncovered GPCR transactivation mediated by adhesion-class GPCRs (aGPCRs). These aGPCR-GPCR transactivation processes regulate unique cell type-specific functions, offering an exciting opportunity to develop therapies that precisely modulate specific GPCR-mediated biological effects.

Noncanonical RGS14 structural determinants control hormone-sensitive NPT2A-mediated phosphate transport.

The sodium phosphate cotransporter-2A (NPT2A) mediates basal and parathyroid hormone (PTH)- and fibroblast growth factor-23 (FGF23)-regulated phosphate transport in proximal tubule cells of the kidney. Both basal and hormone-sensitive transport require sodium hydrogen exchanger regulatory factor-1 (NHERF1), a scaffold protein with tandem PDZ domains, PDZ1 and PDZ2. NPT2A binds to PDZ1. RGS14 persistently represses hormone action by binding to PDZ2. The RGS14 canonical RGS domain, Ras/Rap-binding domains, and G protein regulatory motif cannot explain its regulatory effects on hormone-sensitive phosphate transport because these actions are mediated not only by the PTH receptor, a G protein-coupled receptor (GPCR), but also by the fibroblast growth factor receptor-1, a receptor tyrosine kinase that is not governed by G protein activity. Here, we identify the structural elements of RGS14 that mutually control the action of PTH and FGF23. RGS14 truncation constructs lacking upstream sequence and the RGS domain were fully functional. Removing the linker sequence between the RGS and RBD1 domains abolished RGS14 action. Examination of the alpha-helical linker region suggested candidate serine residues that might facilitate regulatory activities. RGS14 Ser266 and Ser269 are phosphorylated in response to PTH and FGF23, and replacement of these residues by Ala eliminated the actions of RGS14 on hormone-sensitive phosphate transport. PTH stimulated the phosphorylation of a peptide construct harboring the sites of purported phosphorylation and full-length RGS14. Mutating Ser266Ala and Ser269Ala abolished phosphorylation. The results establish that RGS14 regulation of phosphate transport requires targeted phosphorylation within the linker and an intact PDZ ligand.

A Deep Learning Method for Identifying G-Protein Coupled Receptors based on a Feature Pyramid Network and Attention Mechanism

A Deep Learning Method for Identifying G-Protein Coupled Receptors based on a Feature Pyramid Network and Attention Mechanism

Molecular mechanisms of inverse agonism via κ-opioid receptor-G protein complexes.

Opioid receptors, a subfamily of G protein-coupled receptors (GPCRs), are key therapeutic targets. In the canonical GPCR activation model, agonist binding is required for receptor-G protein complex formation, while antagonists prevent G protein coupling. However, many GPCRs exhibit basal activity, allowing G protein association without an agonist. The pharmacological impact of agonist-free receptor-G protein complexes is poorly understood. Here we present biochemical evidence that certain κ-opioid receptor (KOR) inverse agonists can act via KOR-Gi protein complexes. To investigate this phenomenon, we determined cryo-EM structures of KOR-Gi protein complexes with three inverse agonists: JDTic, norBNI and GB18, corresponding to structures of inverse agonist-bound GPCR-G protein complexes. Remarkably, the orthosteric binding pocket resembles the G protein-free 'inactive' receptor conformation, while the receptor remains coupled to the G protein. In summary, our work challenges the canonical model of receptor antagonism and offers crucial insights into GPCR pharmacology.

Targeting cryptic allosteric sites of G protein-coupled receptors as a novel strategy for biased drug discovery.

G protein-coupled receptors (GPCRs) represent the largest family of membrane receptors and are highly effective targets for therapeutic drugs. GPCRs couple different downstream effectors, including G proteins (such as Gi/o, Gs, G12, and Gq) and β-arrestins (such as β-arrestin 1 and β-arrestin 2) to mediate diverse cellular and physiological responses. Biased signaling allows for the specific activation of certain pathways from the full range of receptors' signaling capabilities. Targeting more variable allosteric sites, which are spatially different from the highly conserved orthosteric sites, represents a novel approach in biased GPCR drug discovery, leading to innovative strategies for targeting GPCRs. Notably, the emergence of cryptic allosteric sites on GPCRs has expanded the repertoire of available drug targets and improved receptor subtype selectivity. Here, we conduct a summary of recent progress in the structural determination of cryptic allosteric sites on GPCRs and elucidate the biased signaling mechanisms induced by allosteric modulators. Additionally, we discuss means to identify cryptic allosteric sites and design biased allosteric modulators based on cryptic allosteric sites through structure-based drug design, which is an advanced pharmacotherapeutic approach for treating GPCR-associated diseases.

Shenghui decoction inhibits neuronal cell apoptosis to improve Alzheimer's disease through the PDE4B/cAMP/CREB signaling pathway.

Shenghui Decoction (SHD) is a frequently utilized traditional Chinese medicine formula in clinical settings for addressing cognitive impairment in elderly individuals. Nevertheless, the precise mechanism by which SHD exerts its effects on the most prevalent form of dementia, Alzheimer's disease (AD), remains to be elucidated. Temperature-induced transgenic C. elegans assess Aβ deposition and toxicity. Behavioral experiments are utilized to assess learning and memory capabilities as well as cognitive impairment in APP/PS1 mice. Immunofluorescence and immunohistochemistry are employed to identify Aβ deposits, while UHPLCOE/MS combine network pharmacology is utilized to characterize chemical composition, predict target and analyze the biological processes and signaling pathways modulated by SHD. Molecular biology methodologies confirm the functionality of regulatory pathways. Molecular docking, molecular dynamic simulations (MD) and ultrafiltration-liquid chromatography/mass spectrometry (LC/MS) are employed for the assessment of the binding interactions between active ingredients of SHD and target proteins. SHD effectively reduced the deposition of Aβ in the head of C. elegans and mitigated its toxicity, as well as improved the learning deficits and cognitive impairment in APP/PS1 mice. Network pharmacology analyses revealed that G protein-coupled receptors (GPCRs) and cell apoptosis are the primary biological processes modulated by SHD, with KEEG results indicating that SHD regulated the cAMP signaling pathway. Subsequent experimental investigations demonstrated that SHD attenuated the loss of neurons in APP/PS1 mice, upregulated the expression of anti-apoptotic protein Bcl-2 and downregulated the expression of pro-apoptotic proteins like cleave-Caspase-3 both in vivo and in vitro. Additionally, SHD decreased intracellular AMP levels while increasing cAMP levels, leading to the phosphorylation of PKA to activate CREB. This process ultimately regulated the expression of Bcl-2, Bdnf, among others, to prevent cell apoptosis and safeguard neurons. Molecular docking, MD, and ultrafiltration-LC/MS revealed that the active constituents of SHD formed stable interactions with the cAMP hydrolysis enzyme phosphodiesterase 4B (PDE4B). SHD regulated the cAMP/CREB signaling pathway to inhibit neuronal cell apoptosis and improve AD. Furthermore, it is worth noting that this mechanism may be associated with the specific and consistent binding of SHD active ingredients to PDE4B, potentially offering promising candidates for drug development aimed at addressing AD.

Leveraging Artificial Intelligence in GPCR Activation Studies: Computational Prediction Methods as Key Drivers of Knowledge.

G protein-coupled receptors (GPCRs) are key molecules involved in cellular signaling and are attractive targets for pharmacological intervention. This chapter is designed to explore the range of algorithms used to predict GPCRs' activation states, while also examining the pharmaceutical implications of these predictions. Our primary objective is to show how artificial intelligence (AI) is key in GPCR research to reveal the intricate dynamics of activation and inactivation processes, shedding light on the complex regulatory mechanisms of this vital protein family. We describe several computational strategies that leverage diverse structural data from the Protein Data Bank, molecular dynamic simulations, or ligand-based methods to predict the activation states of GPCRs. We demonstrate how the integration of AI into GPCR research not only enhances our understanding of their dynamic properties but also presents immense potential for driving pharmaceutical research and development, offering promising new avenues in the search for newer, better therapeutic agents.

Modulating vertebrate physiology by genomic fine-tuning of GPCR functions.

G protein-coupled receptors (GPCRs) play a crucial role as membrane receptors, facilitating the communication of eukaryotic species with their environment and regulating cellular and organ interactions. Consequently, GPCRs hold immense potential in contributing to adaptation to ecological niches and responding to environmental shifts. Comparative analyses of vertebrate genomes reveal patterns of GPCR gene loss, expansion, and signatures of selection. Integrating these genomic data with insights from functional analyses of gene variants enables the interpretation of genotype-phenotype correlations. This review underscores the involvement of GPCRs in adaptive processes, presenting numerous examples of how alterations in GPCR functionality influence vertebrate physiology or, conversely, how environmental changes impact GPCR functions. The findings demonstrate that modifications in GPCR function contribute to adapting to aquatic, arid, and nocturnal habitats, influencing camouflage strategies, and specializing in particular dietary preferences. Furthermore, the adaptability of GPCR functions provides an effective mechanism in facilitating past, recent, or ongoing adaptations in animal domestication and human evolution and should be considered in therapeutic strategies and drug development.

Quantitative approaches for studying G protein-coupled receptor signalling and pharmacology.

G protein-coupled receptor (GPCR) signalling pathways underlie numerous physiological processes, are implicated in many diseases and are major targets for therapeutics. There are more than 800 GPCRs, which together transduce a vast array of extracellular stimuli into a variety of intracellular signals via heterotrimeric G protein activation and multiple downstream effectors. A key challenge in cell biology research and the pharmaceutical industry is developing tools that enable the quantitative investigation of GPCR signalling pathways to gain mechanistic insights into the varied cellular functions and pharmacology of GPCRs. Recent progress in this area has been rapid and extensive. In this Review, we provide a critical overview of these new, state-of-the-art approaches to investigate GPCR signalling pathways. These include novel sensors, Förster or bioluminescence resonance energy transfer assays, libraries of tagged G proteins and transcriptional reporters. These approaches enable improved quantitative studies of different stages of GPCR signalling, including GPCR activation, G protein activation, second messenger (cAMP and Ca2+) signalling, β-arrestin recruitment and the internalisation and intracellular trafficking of GPCRs.

FMRFamide G protein-coupled receptors (GPCR) in the cuttlefish Sepiella japonica: Identification, characterization and expression profile

FMRFamide is a ubiquitous neuromodulator in the animal kingdom. Once FMRFamide or similar neuropeptides bind to their G protein-coupled receptors (GPCR), a series of signal transduction events are triggered, thereby mediating various physiological effects. FMRFamide had been reported to be involved in the regulation of sexual maturation in Sepiella japonica. In this research, the full-length cDNA of FMRFamide G protein-coupled receptor of S. japonica (SjFaGPCR) was cloned. The sequence is 1396 bp long and encodes a protein consisting of 418 amino acid residues, lacking a signal peptide at the N-terminal region. The 3D structure of SjFaGPCR was predicted using Todarodes pacificus rhodopsin as a template, and the result indicated the presence of seven transmembrane regions. Multiple sequence alignments and phylogenetic trees indicated that SjFaGPCR is conserved among invertebrates, and shares highly similar sequence characteristics with other cephalopods. In situ hybridization (ISH) results revealed that significant signals of SjFaGPCR were detected in the central medulla and the granular layer cells of the optic lobe, and were also observed in the supraesophageal and subesophageal masses of the brain. Meanwhile, quantitative real-time PCR (qRT-PCR) results showed that a higher expression level of SjFaGPCR mRNA was detected in the brain and optic lobe of female cuttlefish at stage III and stage VI, and also in the brain (stage V) and optic lobe (stages IV and V) of male cuttlefish than that in other tissues. The co-localization results demonstrated that fluorescence signals of SjFMRFamide and SjFaGPCR were overlapped in HEK293 cells, suggesting a possible interaction between the SjFMRFamide and SjFaGPCR. These findings provide molecular support for further exploring the roles of FMRFamide and FaGPCR in the reproductive regulation of S. japonica.

Thermodynamic Role of Receptor Phosphorylation Barcode in Cannabinoid Receptor Desensitization

The endocannabinoid signaling system is comprised of CB1 and CB2 G protein-coupled receptors (GPCRs). CB2 receptor subtype is predominantly expressed in the immune cells and signals through its transducer proteins (Gi protein and β-arrestin-2). Arrestins are signaling proteins that bind to many GPCRs after receptor phosphorylation to terminate G protein signaling (desensitization) and to initiate specific G protein-independent arrestin-mediated signaling pathways via a “phosphorylation barcode”, that captures sequence patterns of phosphorylated Ser/Thr residues in the receptor’s intracellular domains and can lead to different signaling effects. The structural basis for how arrestins and G proteins compete with the receptor for biased signaling and how different barcodes lead to different signaling profiles is not well understood as there is a lack of phosphorylated receptor structures in complex with arrestins. In this work, structural models of β-arrestin-2 were built in complex with the phosphorylated and unphosphorylated forms of the CB2 receptor. The complex structures were relaxed in the lipid bilayer environment with molecular dynamics (MD) simulations and analyzed structurally and thermodynamically. The β-arrestin-2 complex with the phosphorylated receptor was more stable than the non-phosphorylated one, highlighting the thermodynamic role of the receptor phosphorylation. It was also more stable than any of the G protein complexes with CB2 suggesting that phosphorylation signals receptor desensitization (end of G protein signaling) and arrest of the receptor by arrestins. These models are beginning to provide the thermodynamic landscape of CB2 signaling, which can help bias signaling towards therapeutically beneficial pathways in drug discovery applications.

Chemogenetic engagement of different GPCR signaling pathways segregates the orexigenic activity from the control of whole-body glucose metabolism by AGRP Neurons.

BackgroundThe control of energy balance involves neural circuits in the central nervous system, including AGRP neurons in the arcuate nucleus of the hypothalamus (ARC). AGRP neurons are crucial for energy balance and their increased activity during fasting is critical to promote feeding behavior. The activity of these neurons is influenced by multiple signals including those acting on G-protein coupled receptors (GPCR) activating different intracellular signaling pathways. We sought to determine whether discrete G-protein mediated signaling in AGRP neurons, promotes differential regulation of feeding and whole-body glucose homeostasis. Methodology: To test the contribution of Gαq/11 or Gαs signaling, we developed congenital mouse lines expressing the different DREADD receptors (i.e., hM3q and rM3s), in AGRP neurons. Then we elicited chemogenetic activation of AGRP neurons in these mice during the postprandial state to determine the impact on feeding and glucose homeostasis. ResultsActivation of AGRP neurons via hM3q and rM3s promoted hyperphagia. In contrast, only hM3q activation of AGRP neurons of the hypothalamic arcuate nucleus during the postprandial state enhanced whole-body glucose disposal by reducing sympathetic nervous system activity to the pancreas and liver, promoting glucose-stimulated insulin secretion, glycogen deposition and improving glucose tolerance. ConclusionThese data indicate that AGRP neurons regulate food intake and glucose homeostasis through distinct GPCR-dependent signaling pathways and suggest that the transient increase in AGRP neuron activity may contribute to the beneficial effects of fasting on glycemic control.