We have developed an array of assays for nicotinic acetylcholine receptor b
inding and function. [I-125]alpha-Bungarotoxin-, (-)-[H-3]nicotine-, and [H
-3]epibatidine-binding nicotinic acetylcholine receptors were assayed in mo
use brain membranes and sections. Nicotinic acetylcholine receptor function
was quantified using synaptosomal [H-3]dopamine, [H-3]gamma-aminobutyric a
cid ([H-3]GABA), and Rb-86(+) efflux techniques. Additionally, the effects
of beta 2 subunit deletion on each of the measures were assessed. Detailed
pharmacological comparison revealed minimally six nicotinic binding subtype
s: [I-125]alpha-bungarotoxin-binding nicotinic acetylcholine receptors; bet
a 2-subunit-dependent and -independent high-affinity(-)-[H-3]nicotine-bindi
ng sites; beta 2-dependent and -independent cytisine-resistant [H-3]epibati
dine-binding sites; and a beta 2-dependent low-affinity [H-3]epibatidine bi
nding site. Comparative pharmacology suggested that [H-3]GABA and dihydro-b
eta-erythroidine (DH beta E)-sensitive Rb-86(+) efflux are mediated by the
same (probably alpha 4 beta 2) nicotinic acetylcholine receptor subtype, wh
ile other nicotinic acetylcholine receptor subtypes evoke [H-3]dopamine and
DH beta E-resistant Rb-86(+) efflux. In whole-brain preparations, each mea
sure of nicotinic acetylcholine receptor function was beta 2 dependent. The
majority of beta 2-independent [H-3]epibatidine binding was located in sma
ll, scattered brain nuclei, suggesting that individual nuclei may prove sui
table for identification of novel, native nicotinic acetylcholine receptors
. (C) 2000 Elsevier Science B.V. All rights reserved.