Detection of multiple H 3 receptor affinity states utilizing [ 3 H]A‐349821, a novel, selective, non‐imidazole histamine H 3 receptor inverse agonist radioligand

1 A‐349821 is a selective histamine H 3 receptor antagonist/inverse agonist. Herein, binding of the novel non‐imidazole H 3 receptor radioligand [ 3 H]A‐349821 to membranes expressing native or recombinant H 3 receptors from rat or human sources was characterized and compared with the binding of the agonist [ 3 H]N‐ α ‐methylhistamine ([ 3 H]N α MH). 2 [ 3 H]A‐349821 bound with high affinity and specificity to an apparent single class of saturable sites and recognized human H 3 receptors with 10‐fold higher affinity compared to rat H 3 receptors. [ 3 H]A‐349821 detected larger populations of receptors compared to [ 3 H]N α MH. 3 Displacement of [ 3 H]A‐349821 binding by H 3 receptor antagonists/inverse agonists was monophasic, suggesting recognition of a single binding site, while that of H 3 receptor agonists was biphasic, suggesting recognition of both high‐ and low‐affinity H 3 receptor sites. 4 p K i values of high‐affinity binding sites for H 3 receptor competitors utilizing [ 3 H]A‐349821 were highly correlated with p K i values obtained with [ 3 H]N α MH, consistent with labelling of H 3 receptors by [ 3 H]A‐349821. 5 Unlike assays utilizing [ 3 H]N α MH, addition of GDP had no effect on saturation parameters measured with [ 3 H]A‐349821, while displacement of [ 3 H]A‐349821 binding by the H 3 receptor agonist histamine was sensitive to GDP. 6 In conclusion, [ 3 H]A‐349821 labels interconvertible high‐ and low‐affinity states of the H 3 receptor, and displays improved selectivity over imidazole‐containing H 3 receptor antagonist radioligands. [ 3 H]A‐349821 competition studies showed significant differences in the proportions and potencies of high‐ and low‐affinity sites across species, providing new information about the fundamental pharmacological nature of H 3 receptors.British Journal of Pharmacology (2006) 148 , 657–670. doi: 10.1038/sj.bjp.0706752