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

1 We have investigated the binding of a novel histamine H-3-receptor antago nist radioligand, [H-3]-clobenpropit ([H-3]-VUF9153), to guinea-pig cerebra l cortex membranes. 2 Saturation isotherms for [H-3]-clobenpropit appeared biphasic. Scatchard plots were curvilinear and Hill plot slopes were significantly less than un ity (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 (pK(D)') of 10.91 +/- 0.12 (B-max = 5.34 +/- 0.85 fmol mg(-1) original wet weight) and 9.17 +/- 0.16 (B-max = 23.20 +/- 6.70 fmol mg(-1)). 3 In the presence of metyrapone (3 mM) or sodium chloride (100 mM), [H-3]-c lobenpropit appeared to label a homogeneous receptor population (B-max = 3. 41 +/- 0.46 fmol mg(-1) and 3.49 +/- 0.44 fmol mg(-1), pK(D)' = 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 mu M), rilmenidine (3 mu M), idazo xan (0.3 mu M), pentazocine (3 mu M) and 1,3-di-(2-tolyl)guanidine (0.3 mu M) had no effect on the binding of [H-3]-clobenpropit. 4 The specific binding of [H-3]-clobenpropit appeared to reach equilibrium after 25 min at 21 +/- 3 degrees C and remained constant for > 180 min. The estimated pK(D)' (10.27 +/- 0.27; n = 3 +/- s.e.mean) was not significantl y different from that estimated by saturation analysis in the presence of m etyrapone. 5 A series of histamine H-3-receptor ligands expressed affinity values for sites labelled with [H-3]-clobenpropit which were not significantly differe nt from those estimated when [H-3]-R-alpha-MH was used to label histamine H -3-receptors in guinea-pig cerebral cortex membranes.