PT  - JOURNAL ARTICLE
AU  - Thompson, A. J.
AU  - Duke, R. K.
AU  - Lummis, S. C. R.
TI  - Binding Sites for Bilobalide, Diltiazem, Ginkgolide, and Picrotoxinin at the 5-HT&lt;sub&gt;3&lt;/sub&gt; Receptor
AID  - 10.1124/mol.111.071415
DP  - 2011 Jul 01
TA  - Molecular Pharmacology
PG  - 183--190
VI  - 80
IP  - 1
4099  - http://molpharm.aspetjournals.org/content/80/1/183.short
4100  - http://molpharm.aspetjournals.org/content/80/1/183.full
SO  - Mol Pharmacol2011 Jul 01; 80
AB  - Bilobalide (BB), ginkgolide B (GB), diltiazem (DTZ), and picrotoxinin (PXN) are 5-hydroxytryptamine type 3 (5-HT3) receptor antagonists in which the principal sites of action are in the channel. To probe their exact binding locations, 5-HT3 receptors with substitutions in their pore lining residues were constructed (N−4′Q, E−1′D, S2′A, T6′S, L7′T, L9′V, S12′A, I16′V, D20′E), expressed in Xenopus laevis oocytes, and the effects of the compounds on 5-HT-induced currents were examined. EC50 values at mutant receptors were less than 6-fold different from those of wild type, indicating that the mutations were well tolerated. BB, GB, DTZ, and PXN had pIC50 values of 3.33, 3.14, 4.67, and 4.97, respectively. Inhibition by BB and GB was abolished in mutant receptors containing T6′S and S12′A substitutions, but their potencies were enhanced (42- and 125-fold, respectively) in S2′A mutant receptors. S2′A substitution also caused GB ligand trap. PXN potency was modestly enhanced (5-fold) in S2′A, abolished in T6′S, and reduced in L9′V (40-fold) and S12′A (7-fold) receptors. DTZ potency was reduced in L7′T and S12′A receptors (5-fold), and DTZ also displaced [3H]granisetron binding, indicating mixed competitive/noncompetitive inhibition. We conclude that regions close to the hydrophobic gate of M2 are important for the inhibitory effects of BB, GB, DTZ, and PXN at the 5-HT3 receptor; for BB, GB, and PXN, the data show that the 6′ channel lining residue is their major site of action, with minor roles for 2′, 9′, and 12′ residues, whereas for DTZ, the 7′ and 12′ sites are important.