Characterization of the binding of [3H]-clobenpropit to histamine H3-receptors in guinea-pig cerebral cortex membranes

We have investigated the binding of a novel histamine H3-receptor antagonist radioligand, [3H]- clobenpropit ([3H]-VUF9153), to guinea-pig cerebral cortex membranes. Saturation isotherms for [3H]-clobenpropit appeared biphasic. Scatchard plots were curvilinear and Hill plot slopes were significantly less than unity (0.63±0.03; n=12±s.e.mean). The radioligand appeared to label two sites in guinea-pig cerebral cortex membranes with apparent affinities (pKD′) of 10.91±0.12 (Bmax=5.34±0.85 fmol mg−1 original wet weight) and 9.17±0.16 (Bmax=23.20± 6.70 fmol mg−1). In the presence of metyrapone (3 mM) or sodium chloride (100 mM), [3H]-clobenpropit appeared to label a homogeneous receptor population (Bmax=3.41±0.46 fmol mg−1 and 3.49±0.44 fmol mg−1, pKD′ =10.59±0.17 and 10.77±0.02, respectively). Scatchard plots were linear and Hill slopes were not significantly different from unity (0.91±0.04 and 0.99±0.02, respectively). Granisetron (1 μM), rilmenidine (3 μM), idazoxan (0.3 μM), pentazocine (3 μM) and 1,3-di-(2-tolyl)guanidine (0.3 μM) had no effect on the binding of [3H]-clobenpropit. The specific binding of [3H]-clobenpropit appeared to reach equilibrium after 25 min at 21±3°C and remained constant for >180 min. The estimated pKD′ (10.27±0.27; n=3±s.e.mean) was not significantly different from that estimated by saturation analysis in the presence of metyrapone. A series of histamine H3-receptor ligands expressed affinity values for sites labelled with [3H]-clobenpropit which were not significantly different from those estimated when [3H]-R-α-MH was used to label histamine H3-receptors in guinea-pig cerebral cortex membranes. British Journal of Pharmacology (1999) 128, 881–890; doi:10.1038/sj.bjp.0702860