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Investigating the ligand agonism and antagonism at the D_2long receptor by dynamic mass redistribution.

ABSTRACT: The signalling of the D₂ receptor (D₂R), a G protein-coupled receptor (GPCR), is a complex process consisting of various components. For the screening of D₂R ligands, methods quantifying distinct second messengers such as cAMP or the interaction of the receptor with β-arrestin, are commonly employed. In contrast, a label-free biosensor technology like dynamic mass redistribution (DMR), where it is mostly unknown how the individual signalling pathways contribute to the DMR signal, provides a holistic readout of the complex cellular response. In this study, we report the successful application of the DMR technology to CHO-K1 cells stably expressing the human dopamine D_2long receptor. In real-time kinetic experiments, studies of D₂R reference compounds yielded results for agonists and antagonists that were consistent with those obtained by conventional methods and also allowed a discrimination between partial and full agonists. Furthermore, investigations on the signalling pathway in CHO-K1 hD_2longR cells identified the Gα_i/o protein as the main proximal trigger of the observed DMR response. The present study has shown that the DMR technology is a valuable method for the characterisation of putative new ligands and, due to its label-free nature, suggests its use for deorphanisation studies of GPCRs.

SUBMITTER: Forster L

PROVIDER: S-EPMC9187652 | biostudies-literature | 2022 Jun

REPOSITORIES: biostudies-literature

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Investigating the ligand agonism and antagonism at the D2long receptor by dynamic mass redistribution.

Forster Lisa L Pockes Steffen S

Scientific reports 20220610 1

The signalling of the D2 receptor (D2R), a G protein-coupled receptor (GPCR), is a complex process consisting of various components. For the screening of D2R ligands, methods quantifying distinct second messengers such as cAMP or the interaction of the receptor with β-arrestin, are commonly employed. In contrast, a label-free biosensor technology like dynamic mass redistribution (DMR), where it is mostly unknown how the individual signalling pathways contribute t ...[more]

PMID: 35688965

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Project description:ObjectiveTreatment with glucagon receptor antagonists (GRAs) reduces blood glucose but causes dyslipidemia and accumulation of fat in the liver. We investigated the acute and chronic effects of glucagon on lipid metabolism in mice.MethodsChronic effects of glucagon receptor signaling on lipid metabolism were studied using oral lipid tolerance tests (OLTTs) in overnight fasted glucagon receptor knockout (Gcgr-/-) mice, and in C57Bl/6JRj mice treated with a glucagon receptor antibody (GCGR Ab) or a long-acting glucagon analogue (GCGA) for eight weeks. Following treatment, liver tissue was harvested for RNA-sequencing and triglyceride measurements. Acute effects were studied in C57Bl/6JRj mice treated with a GRA or GCGA 1 h or immediately before OLTTs, respectively. Direct effects of glucagon on hepatic lipolysis were studied using isolated perfused mouse liver preparations. To investigate potential effects of GCGA and GRA on gastric emptying, paracetamol was, in separate experiments, administered immediately before OLTTs.ResultsPlasma triglyceride concentrations increased 2-fold in Gcgr-/- mice compared to their wild-type littermates during the OLTT (P = 0.001). Chronic treatment with GCGR Ab increased, whereas GCGA treatment decreased, plasma triglyceride concentrations during OLTTs (P < 0.05). Genes involved in lipid metabolism were upregulated upon GCGR Ab treatment while GCGA treatment had opposite effects. Acute GRA and GCGA treatment, respectively, increased (P = 0.02) and decreased (P = 0.003) plasma triglyceride concentrations during OLTTs. Glucagon stimulated hepatic lipolysis, evident by an increase in free fatty acid concentrations in the effluent from perfused mouse livers. In line with this, GCGR Ab treatment increased, while GCGA treatment decreased, liver triglyceride concentrations. The effects of glucagon appeared independent of changes in gastric emptying of paracetamol.ConclusionsGlucagon receptor signaling regulates triglyceride metabolism, both chronically and acutely, in mice. These data expand glucagon´s biological role and implicate that intact glucagon signaling is important for lipid metabolism. Glucagon agonism may have beneficial effects on hepatic and peripheral triglyceride metabolism.

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Investigating the ligand agonism and antagonism at the D_2long receptor by dynamic mass redistribution.

Publications

Investigating the ligand agonism and antagonism at the D<sub>2long</sub> receptor by dynamic mass redistribution.

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