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SB‐258585 (4‐Iodo‐N‐[4‐methoxy‐3‐(4‐methyl‐piperazin‐1‐yl)‐phenyl]‐benzenesulphonamide) is a high affinity ligand at 5‐HT6 receptors. It displays over 100 fold selectivity for the 5‐HT6 receptor over all other 5‐HT receptors tested so far. SB‐258585 has been radiolabelled, to high specific activity, for its characterization as a 5‐HT6 receptor selective radioligand. [125I]‐SB‐258585 bound, with high affinity, to a single population of receptors in a cell line expressing human recombinant 5‐HT6 receptors. Kinetic and saturation binding experiments gave pKD values of 9.01±0.09 and 9.09±0.02, respectively. In membranes derived from rat or pig striatum and human caudate putamen, [125I]‐SB‐258585 labelled a single site with high levels (>60%) of specific binding. Saturation analysis revealed pKD values of 8.56±0.07 for rat, 8.60±0.10 for pig and 8.90±0.02 for human. Bmax values for the tissues ranged from 173±23 and 181±25 fmol mg−1 protein in rat and pig striatum, respectively, to 215±41 fmol mg−1 protein in human caudate putamen. The pKi rank order of potency for a number of compounds, determined in competition binding assays with [125I]‐SB‐258585, at human caudate putamen membranes was: SB‐271046>SB‐258585>SB‐214111>methiothepin>clozapine>5‐Me‐OT>5‐HT>Ro 04‐6790>mianserin>ritanserin=amitriptyline>5‐CT>mesulergine. Similar profiles were obtained from pig and rat striatal membranes and recombinant 5‐HT6 receptors; data from the latter correlated well with [3H]‐LSD binding. Thus, [125I]‐SB‐258585 is a high affinity, selective radioligand which can be used to label both recombinant and native 5‐HT6 receptors and will facilitate further characterization of this receptor subtype in animal and human tissues. British Journal of Pharmacology (2000) 130, 1597–1605; doi:

3 In functional studies on human 5-HT 6 receptors SB-271046 competitively antagonized 5-HTinduced stimulation of adenylyl cyclase activity with a pA 2 of 8.71. 4 SB-271046 produced an increase in seizure threshold over a wide-dose range in the rat maximal electroshock seizure threshold (MEST) test, with a minimum e ective dose of 40.1 mg kg 71 p.o. and maximum e ect at 4 h post-dose. The level of anticonvulsant activity achieved correlated well with the blood concentrations of SB-271046 (EC 50 of 0.16 mM) and brain concentrations of 0.01 ± 0.04 mM at C max . 5 These data, together with the observed anticonvulsant activity of other selective 5-HT 6 receptor antagonists, SB-258510 (10 mg kg 71 , 2 ± 6 h pre-test) and Ro 04-6790 (1 ± 30 mg kg 71 , 1 h pre-test), in the rat MEST test, suggest that the anticonvulsant properties of SB-271046 are likely to be mediated by 5-HT 6 receptors. 6 Overall, these studies demonstrate that SB-271046 is a potent and selective 5-HT 6 receptor antagonist and is orally active in the rat MEST test. SB-271046 represents a valuable tool for evaluating the in vivo central function of 5-HT 6 receptors.