Stock_2014_J.Physiol_592_2501

'Embryonic' muscle-type nicotinic acetylcholine receptor channels (nAChRs) bind ligands at interfaces of alpha- and gamma- or delta-subunits. alphagamma and alphadelta sites differ in affinity, but their contributions to opening the channel have remained elusive. We compared high-resolution patch clamp currents evoked by epibatidine (Ebd), carbamylcholine (CCh) and acetylcholine (ACh). Ebd binds with 75-fold higher affinity at alphagamma than at alphadelta sites, whereas CCh and ACh prefer alphadelta sites. Similar short (tauO1), intermediate (tauO2) and long (tauO3) types of opening were observed with all three agonists. tauO2 openings were maximally prevalent at low Ebd concentrations, binding at alphagamma sites. By contrast, tauO1 openings appear to be generated at alphadelta sites. In addition, two types of burst appeared: short bursts of an average of 0.75 ms (tauB1) that should arise from the alphagamma site, and long bursts of 12-25 ms (tauB2) in duration arising from double liganded receptors. Limited by the temporal resolution, the closings within bursts were invariant at 3 mus. Corrected for missed closings, in the case of ACh the openings within long bursts lasted 170 mus and those in short bursts about 30 mus. Blocking alphadelta sites with alpha-conotoxin M1 (CTx) eliminated both tauO1 and tauB2 and left only tauO2 and the short tauB1 bursts, as expected. Furthermore we found desensitization when the receptors bound ACh only at the alphagamma site. When CTx was applied to 'embryonic' mouse endplates, monoquantal current rise times were increased, and amplitude and decay time constants were reduced, as expected. Thus the alphagamma and alphadelta sites of nAChRs elicit specific channel-opening patterns.