Characterisation of the binding of [3H]methyllycaconitine:: a new radioligand for labelling α7-type neuronal nicotinic acetylcholine receptors

Methyllycaconitine (MLA), a norditerpenoid alkaloid isolated from Delphinium seeds, is one of the most potent non-proteinacious ligands that is selective for abungarotoxin-sensitive neuronal nicotinic acetylcholine receptors (nAChR). [H-3]MLA bound to rat brain membranes with high affinity (K-d = 1.86 +/- 0.31 nM) with a good ratio of specific to non-specific binding. The binding of [H-3]MLA was characterised by rapid association (t(1/2) = 2.3 min) and dissociation (t(1/2) = 12.6 min) kinetics. The radioligand binding displayed nicotinic pharmacology, consistent with an interaction with alpha bungarotoxin-sensitive nAChR. The snake alpha-toxins, alpha bungarotoxin and alpha cobratoxin, displaced [H-3]MLA with high affinity (K-i = 1.8 +/- 0.5 and 5.5 +/- 0.9 nM, respectively), whereas nicotine was less potent (K-i = 6.1 +/- 1.1 mu M). The distribution of [H-3]MLA binding sites in crudely dissected rat brain regions was identical to that of [I-125]alpha bungarotoxin binding sites, with a high binding site density in hippocampus and hypothalamus, but low density in striatum and cerebellum. [H-3]MLA also labelled a sub-population of binding sites which are not sensitive to the snake alpha toxins, but which did not differ significantly from the major population with respect to their other pharmacological properties or regional distribution. [H-3]MLA, therefore, is a novel radiolabel for characterising alpha 7-type nAChR. A good signal to noise ratio and rapid binding kinetics provide advantages over the use of radiolabelled alpha bungarotoxin for rapid and accurate equilibrium binding assays. (C) 1999 Elsevier Science Ltd. All rights reserved.