Effects of Redox Reagents and Arsenical Compounds on [3H]‐Cytisine Binding to Immunoisolated Nicotinic Acetylcholine Receptors from Chick Brain Containing α4 β2 Subunits

All known nicotinic receptor α subunits include a conserved disulfide bond that is essential for function and is a site for labeling via biochemical modification. In an effort to develop a universal ligand for all subtypes of nicotinic receptors, we previously studied the effects of arsenylation with two compounds, ρ‐aminophenyldichloroarsine (APA) and bromoacetyl‐ρ‐aminophenylarsenòxide (BAPA) on nicotinic receptors from Torpedo electroplax. Here we apply these reagents to immunoisolated receptors containing α4, β2, and possibly other subunits from chick brain that bind [3H]cytisine with high affinity (KD∼5 nM). These are distinct from another receptor subtype that also binds [3H]cytisine and [3H]nicotine and can be arsenylated with APA, but instead contains α5,β2, and probably other subunits. Reduction of α4 β2 receptors with dithiothreitol blocked [3H]cytisine binding and this effect was reversed upon reoxidation by dithiobisnitrobenzoic acid. APA or BAPA prevented the dithiobisnitrobenzoic acid reactivation of dithiothreitol‐treated receptors with IC50 values of 15 and 70 nM, respectively. However, the antiarsenical dimercaptopropanesulfonic acid restored function to APA‐ or BAPA‐ “arsenylated” receptors (EC50∼100 μM). APA‐treated receptors remained blocked for up to 24 h, but treatment with dimercaptopropanesulfonic acid at any time restored [3H]cytisine binding. APA treatment of reduced receptors protected against irreversible alkylation by Bromoacetyl choline, indicating that arsenylation occurs at least in part in the agonist binding site. Thus, these reagents have similar effects on different nicotinic receptor subtypes from both muscle and nerves.