AAV ACTOne Gs-GPCR Assay Kit-AMY3

The AAV ACTOne Gs-GPCR Assay Kit-AMY3 is designed specifically for screening drugs/compounds that target the Amylin Receptor 3 (AMY3) by combination AAV and ACTOne technologies.
Amylin Receptor 3 (AMY3) is a receptor complex that consists of the calcitonin receptor (CTR) and receptor activity-modifying protein 3 (RAMP3). This receptor is part of the amylin receptor family and is activated by the hormone amylin, which is co-secreted with insulin by pancreatic beta cells. AMY3 is involved in regulating glucose metabolism, appetite, and gastric emptying. It plays a role in the central nervous system's control of energy homeostasis and is being explored as a target for treating metabolic disorders such as obesity and diabetes.
This kit uses AAV vectors with a CMV promoter to co-express the AMY3 and cyclic nucleotide-gated (CNG) channel, allowing researchers to conduct high-throughput screening and functional analysis of potential AMY3-targeting compounds. The kit provides a sensitive and reliable method for evaluating the pharmacological properties of AMY3 drugs, such as agonists and antagonists, in a live-cell environment.

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Background

ACTOne™ is the only high-throughput GPCR screening technology that can directly measure the intracellular changes of the secondary messenger cyclic AMP (cAMP) in living cells, in real-time. It uses a proprietary modified cyclic nucleotide-gated (CNG) channel, which is co-localized with adenylate cyclase at the plasma membrane, as a biosensor of cAMP activity. The CNG channel opens when the cAMP level near the plasma membrane increases, resulting in ion flux and cell membrane depolarization. The influx of cations through the CNG channel can be quantified using fluorescent ion indicators or membrane potential (MP) dyes. It provides information on real time intracellular cAMP changes and is highly sensitive. By combining kinetic and endpoint readouts, we are able to capture and analyze transient responses from endogenous GPCRs and weak responses caused by weak Gs or Gi coupled GPCR activities. Using ACTOne, we are able to detect the subcellular cAMP concentration changes directly caused by GPCR activation. Real-time kinetic readouts minimize artifacts, and provide greater content and more statistically relevant data. The intensity of signal increase caused by GPCR activation is directly related to the receptor number on cell surface. Using ACTOne assay, we were able to detect activities of some endogenous Gs coupled receptors in HEK293 cells that have not been reported in literature. In addition, we have also detected weak Gs coupled activity of a GPCR that was widely considered to be only linked to Gq coupled pathway. The ACTOne assay also provides a useful tool for GPCR de-orphanization.