Functional screening of adrenergic receptors by measuring intracellular calcium using the FlexStation scanning fluorimeter

Abstract

In this study we test whether functional screening of compounds to adrenergic G protein-coupled receptors (GPCRs) would provide data that correlated significantly with radiolabeled binding data, thereby permitting researchers to replace expensive radioligand-binding experiments with non-radioactive screening. An increase in intracellular calcium levels represents an important second messenger signal for several recombinant GPCRs. In this study, we describe the affinities of three alpha adrenoceptor antagonists (terazosin, tamsulosin and alfuzosin), determined by monitoring the changes in intracellular calcium levels and comparing them with their radioligand-binding affinities. In addition to determining the functional affinities of the three alpha adrenoceptor antagonists, we evaluate their binding at two alpha adrenoceptor subtypes and optimized the assay for high-throughput screening.