The 5-hydroxytryptamine, receptor-selective radioligand [3H]Ro 63-0563 labels 5-hydroxytryptamine receptor binding sites in rat and porcine striatum

Ro 63-0563 [4-amino-N-(2,6 bis-methylamino-pyridin-4-yl)benzene sulfonamide] is a high affinity 5-hydroxytryptamine(6) (HT6) receptor antagonist with more than 100-fold selectivity for the 5-HT6 receptor compared with 69 other receptors and binding sites. The present study describes the properties of [H-3]Ro 63-0563, the first selective 5-HT6 receptor radioligand. Specific binding of [H-3]Ro 63-0563 (nonspecific binding defined in the presence of 10 mu M methiothepin) to recombinant rat and human 5-HT6 receptors was saturable, rapid, and reversible with equilibrium dissociation constants (K-d) of 6.8 nM and 4.96 nM, respectively. The pharmacological profile of the rat 5-HT6 receptor labeled with [H-3]Ro 63-0563 (methiothepin > D-lysergic acid diethylamide > clozapine similar to Ro 63-0563 > lisuride > ergotamine similar to Ro 04-6790 > 5-HT > amitriptytine similar to metergoline similar to mianserin similar to ritanserin > methysergide > mesulergine) was similar to that obtained by using either [H-3]D-lysergic acide diethylamide or [H-3]5-HT as radioligand. In equilibrium binding studies with rat striatal membranes, [H-3]Ro 63-0563 labeled a single binding site with K-d and B-max values of 11.7 nM and 175 fmol/mg protein, respectively. In porcine striatal membranes, [H-3]Ro 63-0563 also labeled a single binding site with K-d and B-max values of 8.0 nM and 130 fmol/mg protein, respectively. The affinities of 14 5-HT6 receptor ligands at this binding site were similar to those found for the recombinant rat and human 5-HT6 receptor, which suggested the presence of 5-HT6 receptors in porcine striatum.