High-throughput real-time monitoring of Gs-coupled receptor activation in intact cells using cyclic nucleotide-gated channels

Eur J Pharmacol. 2003 Sep 30;478(1):27-34. doi: 10.1016/j.ejphar.2003.08.036.


Cyclic adenosine-monophosphate (cAMP) is one of the major second messenger molecules transmitting extracellular stimuli into short- and long-term changes of intracellular homeostasis. Measurements of cellular cAMP levels are often used to quantify and characterize signaling by G protein-coupled receptors. Current assays for cAMP determination are usually end-point assays involving cell lysis. We have developed a technology to monitor real-time changes of cAMP levels in living cells. This method uses a modified cyclic nucleotide-gated (CNG) Ca(2+) channel which is opened by intracellular cAMP. Thus, changes in cAMP levels are translated into changes in free Ca(2+) which can easily be measured using fluorimetric imaging technologies compatible with high-throughput screening formats. The new assay method was used to characterize the pharmacology of various endogenously and heterologously expressed G protein-coupled receptors and allows for the simultaneous study of G(s), G(i) and G(q)-linked receptors in the same cell population.

Publication types

  • Comparative Study
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Cell Line
  • Cyclic AMP / agonists
  • Cyclic AMP / metabolism
  • Cyclic Nucleotide-Gated Cation Channels
  • Dose-Response Relationship, Drug
  • Epinephrine / pharmacology
  • GTP-Binding Protein alpha Subunits, Gs / metabolism*
  • Humans
  • Ion Channels / agonists
  • Ion Channels / metabolism*
  • Octopamine / pharmacology
  • Rats
  • Time Factors


  • Cyclic Nucleotide-Gated Cation Channels
  • Ion Channels
  • Octopamine
  • Cyclic AMP
  • GTP-Binding Protein alpha Subunits, Gs
  • Epinephrine