NEURONAL NICOTINIC ALPHA-7 RECEPTOR EXPRESSED IN XENOPUS OOCYTES PRESENTS 5 PUTATIVE BINDING-SITES FOR METHYLLYCACONITINE

1. The recently isolated compound methyllycaconitine (MLA) is a plant toxin which is a competitive inhibitor of nicotinic acetylcholine rece ptors (nAChRs). We found that homomeric alpha 7 receptors display a ve ry high sensitivity to MLA with an IC50 in the picomolar range. 2. The competitive nature of the alpha 7 MLA blockade was reinforced by the observation that this compound has no action on wild-type serotoninerg ic receptors (5-HT3), whereas it is a powerful antagonist of chimaeric receptors alpha 7-5-HT3. 3. The time course of MLA inhibition of the wild-type (WT) alpha 7 follows a monotonic exponential decay whose tim e constant is proportional to the MLA concentration and could be descr ibed by a bimolecular mechanism with a forward rate constant (k(+)) of 2.7 x 10(7) s(-1) M(-1). In contrast, recovery from MLA inhibition di splays an S-shaped time course that is incompatible with a simple bimo lecular reaction. 4. Given the pentameric nature of the neuronal nicot inic receptors, a linear chain model, including five putative MLA bind ing sites corresponding to the homomeric nature of alpha 7, is propose d. 5. Both onset and recovery data obtained on the alpha 7 wild-type r eceptor are adequately described by this model assuming that a single MLA molecule is sufficient to block receptor function. 6. analysis of MLA blockade and recovery of reconstituted heteromeric alpha 4 beta 2 receptors reveals, as expected, a time course compatible with only two binding sites for the toxin and, thus, further supports the validity of our model.