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Gq-coupled Receptor Assays Using FLEXstation

Alexander C. Zambon1, Bruce R. Conklin1, Laurence L. Brunton2,3, Paul A. Insel2,3, and Anne T. Ferguson4

1Gladstone Institute of Cardiovascular Disease, University of California, San Francisco, CA 94141-9100
Departments of Pharmacology2 and Medicine3, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0636
4Molecular Devices Corp., 1311 Orleans Drive, Sunnyvale, CA 94089


Introduction
Gq-coupled receptors are cell-surface proteins that signal through multiple pathways, resulting in the release of Ca2+ from intracellular storage vesicles. They are gaining widespread popularity because they are implicated in critical intra- and intercellular processes. Since abnormal regulation of Gq-coupled receptor signaling pathways can cause various diseases, these receptors are prime targets for drug therapy. For this reason, it is important to study the regulation and specificity of Gq-coupled receptor activity.

Traditionally, Gq-coupled receptor activity is monitored by measuring an increase in intracellular inositol triphosphates (IP3) and/or calcium ions (Ca2+) (Fig. 1). Inositol phosphate measurement using [3H] inositol is labor intensive and requires radioactive waste storage and removal. In comparison, the release of calcium from intracellular stores into the cytoplasm was traditionally monitored in single cells by using fluorescence microscopy image analysis and a calcium-sensitive fluorescent dye. This procedure, while allowing a view of individual cells, is laborious and time-consuming when multiple ligands or drugs are being tested.


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