GPCR Biology
G-Protein Coupled Receptors (GPCRs) are one of the largest receptor families in the genome and are essential for the healthy function of nearly every organ in the body. GPCRs are also important targets for therapeutic drugs. Increase your understanding of drug effects and GPCR biology with bright fluorescent assays in living cells.
With our GPCR Assays in Living Cells, you can:
- Detect kinetic Gs, Gi, and Gq mediated responses in living cells
- Combine multiple assays in the same cell population
- Express sensors and run assays in disease relevant cell types
- Detect fluorescence on imaging systems or automated plate readers (Z’>0.8)
- Quantify agonist bias
- De-orphanize receptors
Detecting Gs
Detect Gs mediated cAMP fluctuations with the cADDis cAMP assay.
→ BMG CLARIOstar® Application Note for Gs
Detecting Gi
Our cADDis cAMP sensor can be also used to detect Gi mediated responses.
→ Request Gi Assay Paper
Detecting Gq
Detect Gq mediated responses using R-GECO Ca2+, DAG, and PIP2 assays.
→ BMG CLARIOstar® Application Note for Gq
Continuous Multiplex Measurements
Red fluorescent DAG sensor multiplexed with cADDis, a green fluorescent cAMP sensor, indicates Gs & Gq signaling via a calcitonin receptor.
Combine our red and green sensors in the same cells to measure multiple analytes simultaneously. Simply add both sensors to the transduction mix. Measure two signals, examine ligand bias, deorphanize GPCRs, and look for off-target GPCR activation.
Assays in Primary Cultures or iPSCs
Detect GPCR mediated responses in cells relevant to your disease, drug target, or biology of interest. Our BacMam-packaged sensors have been used in neurons, cardiomyoctes, islets, and many more primary and iPSC derived cells! Alternate promoters or viral vectors are available by request. The BacMam Transducible Cells page lists some publications showing disease-relevant cells transduced with BacMam. Pictured, at right, are the cADDis cAMP Assay in primary striatal neurons and the Red GECO calcium assay in nCardia’s Cor.4u cardiomyocytes.
Simple Protocol
We strive to create simple protocols with minimal liquid handling. Cell lysis, IBMX, enzymes, and co-factors are not necessary for these assays. Add our sensors to your cells, incubate, add drug, and measure fluorescent changes.
Beta Arrestin Assays
Fluorescent beta-arrestin assays to detect arrestin recruitment at specific GPCRs are available as well. They contain the Arrestin sensor, GPCR and relevant GRK in BacMam, positive control agonist, and sodium butyrate.
Available GPCRs include Angiotensin II (AT1R), Vasopressin (V2R), GLP-1, Mu Opioid (OPRM1), and beta2-adrenergic (B2AR) receptors. We are continuing to validate new receptors, if you do not see your receptor of interest please send us an email!
Quantifying bias is easier than you think. Check out this video from Pharmechanics to learn how.
Sound intriguing? Get more details on each of our specific GPCR Assays
cADDis cAMP Assay
DAG Assay
PIP2 Assay
R-GECO Ca2+ Assay
Recent Publications
- I. Gapallawa, et al. Bitter taste receptors stimulate phagocytosis in human macrophages through calcium, nitric oxide, and cyclic-GMP signaling Cell. Mol. Life Sci. Feb, 2020 (bioRxiv)
- D. McMahon, et al. Neuropeptide regulation of secretion and inflammation in human airway gland serous cells (bioRxiv)
- M. Thomas, et al. Design for Fast Optogenetic Screen in Mammalian Cells For Next Gen Ca2+ Sensors. BioRxiv. Feb 2020.
- Nunez, F.J., Johnstone, T.B. et al. Glucocorticoids rapidly activate cAMP production via Gαs to initiate non‐genomic signaling that contributes to one‐third of their canonical genomic effects FASEB Journal, Dec. 2019
- Nunez, F.J., Schulte, N.A., Fogel, D.M., et al. Agonist-specific desensitization of PGE2-stimulated cAMP signaling due to upregulated phosphodiesterase expression in human lung fibroblasts Naunyn-Schmiedeberg’s Arch Pharmacol Dec. 2019
- K. Harlen, et al. Live-Cell Assays for Cell Stress Responses Reveal New Patterns of Cell Signaling Caused by Mutations in Rhodopsin, α-Synuclein and TDP-43 Front. Cell. Neurosci.,December 2019
- K. Hilgendorf, et al. Omega-3 Fatty Acids Activate Ciliary FFAR4 to Control Adipogenesis Cell November, 2019
- R. Forteza, et al. Glucocorticoids and Myosin5b loss-of-function induce heightened PKA signaling in addition to membrane traffic defects MBoC Oct, 2019
- T. Baldwin, et al. Insights into the Regulatory Properties of Human Adenylyl Cyclase Type 9 Molecular Pharmacology. April, 2019
- T. Togo Autocrine purinergic signaling stimulated by cell membrane disruption is involved in both cell membrane repair and adaptive response in MDCK cells Biochem & Biophysical Research Comm. March, 2019
- C. Ojiaku, et al. Transforming Growth Factor-β1 Decreases β2-Agonist–induced Relaxation in Human Airway Smooth Muscle ATSJournas, Dec 2018
- Y.A.B. Sierra, et al. Potassium channel-based optogenetic silencing. Nature Communications Nov. 5 2018.
- X. Chen, et al. Phenylephrine, a common cold remedy active ingredient, suppresses uterine contractions through cAMP signalling. Scientific Reports Aug. 2018.
- P. Tewson, et al. Assay for Detecting Gαi-Mediated Decreases in cAMP in Living Cells. SLAS. July 10, 2018. Request Full Text PDF
- T. Buranda, et al. A High-Throughput Flow Cytometry Screen Identifies Molecules that Inhibit Hantavirus Cell Entry. SLAS Discovery. April 2, 2018.
- N. H. Wray, et al. NMDAR-independent, cAMP-dependent antidepressant actions of ketamine. Molecular Psychiatry. April 2, 2018.
- H. Zou, et al. PDE8: A Novel Target in Airway Smooth Muscle. American Journal of Respiratory Cell and Molecular Biology. Vol. 58, No. 4. April 01 2018.
- M. Thomas, et al. Toward a High Throughput Screen for Voltage Sensors. 2018.
- T.B. Johnstone, et al. PDE8 is Expressed in Human Airway Smooth Muscle and Selectively Regulates cAMP Signaling by β2AR-AC6. American Journal of Respiratory Cell and Molecular Biology. Dec. 2017.
- K.A. McCrink, et al. β-Arrestin2 Improves Post-Myocardial Infarction Heart Failure via Sarco(endo)plasmic Reticulum Ca2+-ATPase-Dependent Positive Inotropy in Cardiomyocytes. Hypertension. Nov. 2017.
- J. Almaça, et al. Human beta cells produce and release serotonin to inhibit glucagon secretion from alpha cells.Cell Reports, Volume 17, Issue 12, 2016.
- P. Tewson, et al. New DAG and cAMP Sensors Optimized for Live-Cell Assays in Automated Laboratories. J Biomol Screen Dec. 11, 2015. Request Full Text PDF
- B.S. Moore, et al. Cilia have high cAMP levels that are inhibited by Sonic Hedgehog-regulated calcium dynamics. PNAS Nov. 15, 2016.
- C Deisl et al. Hypertrophy of human embryonic stem cell–derived cardiomyocytes supported by positive feedback between Ca2+ and diacylglycerol signals. Pflügers Archiv – European Journal of Physiology Jun. 2019
- Herranz, Gonzalo et al. Protein Kinase C δ Regulates the Depletion of Actin at the Immunological Synapse Required for Polarized Exosome Secretion by T Cells. Frontiers in Immunology Apr. 2019
- Ohno, et al. Dynamics of Presynaptic Diacylglycerol in a Sensory Neuron Encode Differences between Past and Current Stimulus Intensity. Cell Reports, Vol. 20, Issue 10. Sept. 2017
- Shigeto, et al. GLP-1 stimulates insulin secretion by PKC-dependent TRPM4 and TRPM5 activation. J Clinical Invest. 2015. doi:10.1172/JCI81975.
- Xingjuan, et al. PKC-dependent Phosphorylation of the H1 Histamine Receptor Modulates TRPC6 Activity. Cells. 2014.
- P. Tewson, et al. New DAG and cAMP Sensors Optimized for Live-Cell Assays in Automated Laboratories. J Biomol Screen Dec. 2015. Request Full Text PDF
- Arttamangkul, et al.Does PKC activation increase the homologous desensitization of μ opioid receptors?British Journal of Pharmacology April 2014
- Tewson, et al. Simultaneous Detection of Ca2+and Diacylglycerol Signaling in Living Cells. PLoS One. 2012.
- L.Liu, et al. Gαq sensitizes TRPM8 to inhibition by PI(4,5)P2 depletion upon receptor activation. J.Neurosci. May 2019.
- , and Teaching an Old Drug New Tricks: Agonism, Antagonism, and Biased Signaling of Pilocarpine through M3 Muscarinic Acetylcholine Receptor Molecular Pharmacology Nov. 2018
- Q. Wang, et al. Regulator of G protein signaling Gβ5-R7 is a crucial activator of muscarinic M3 receptor-stimulated insulin secretion. FASEB J. Jul. 7,2017.
- Y. Ding, et al. Ratiometric biosensors based on dimerization dependent fluorescent protein exchange. Nature Methods, 2015.
- P. Tewson, et al. A multiplexed fluorescent assay for independent second-messenger systems: decoding GPCR activation in living cells. J. Biomolecular Screening 18, 2013.
- M. Thomas, et al. Design for Fast Optogenetic Screen in Mammalian Cells For Next Gen Ca2+ Sensors. BioRxiv. Feb 2020.
- K. Harlen, et al. Live-Cell Assays for Cell Stress Responses Reveal New Patterns of Cell Signaling Caused by Mutations in Rhodopsin, α-Synuclein and TDP-43 Front. Cell. Neurosci.,December 2019
- M.C. Cañizal, et al. A Dual Ca2+/ DAG Sensor Reports on Ligand Efficiency.
- C. Xingjuan, et al. PKC-dependent Phosphorylation of the H1 Histamine Receptor Modulates TRPC6 Activity. Cells. 2014.
- P. Tewson, et al. A multiplexed fluorescent assay for independent second-messenger systems: decoding GPCR activation in living cells. Journal of Biomolecular Screening 18, 2013.
- J. Wu, et al. Improved Orange and Red Ca2+ Indicators and Photophysical Considerations for Optogenetic Applications. ACS Chem Neurosci. Jun. 19, 2013
- H.J. Carlson, et al. Mutational Analysis of a Red Fluorescent Protein-Based Calcium Ion Indicator. Sensors, 2013.
- P. Tewson, et al. Simultaneous Detection of Ca2+ and Diacylglycerol Signaling in Living Cells. PLoS One. 2012.
- Characterizing GPCR Activaton using Live Cell Imaging. Biotek Instruments
- Real-Time Detection of Gs and Gi Signaling in Living Cells on a BMG CLARIOstar. BMG LabTech
- Simultaneous detection of GPCR second messengers in living cells. BMG LabTech
- Thapsigargin-induced Cellular Stress Response and Inhibition of Gq-dependent Calcium Signaling. Biotek Instruments
