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Under normal conditions, individual nerve cells express a cohort of several nicotinic acetylcholine receptor (nAChR) subtypes with unique functional properties. Therefore, single- channel recordings of such neurons reflect the mixed properties of the various expressed receptors.Previous attempts to recapitulate the properties of naturally occurring receptors using recombinant receptors of known subunit composition in heterologous expression systems have been largely unsuccessful, as the properties of these receptors vary widely among expression systems.We measured the properties of specific nAChRs in superior cervical ganglion neurons cultured from mice with targeted deletions of select nAChR subunit genes. Mice lacking both the α5 and the β2 subunits express α3β4 receptors, whereas single-knockout (KO) mice lacking either α5 or β2 express α3β4β2 and α3β4α5 (plus α3β4) hetero-oligomeric receptors, respectively. This approach allows one to investigate these receptors at the single-channel level in their native environment. The single-channel properties of nACh receptors in superior cervical ganglion (SCG) neurons from α5β2α7 triple-KO mice were similar to the properties of receptors measured in α5β2 double-KO mice.The principal conductance level of α3β4 receptors was 32.6 ± 0.8 pS (mean ± SEM), and these receptors also displayed both higher and lower secondary conductance levels. The conductance levels of α3β4α5 receptors were identical to α3β4 receptors, but the α3β4α5 receptors had longer open times and burst duration. α3β4β2 receptors had a lower conductance level and longer open times than α3β4 receptors. Interestingly, all three receptor types could be identified faithfully in wild-type C57Bl/6J SCG neurons.Previous attempts to measure the functional properties of recombinant nicotinic acetylcholine receptors (nAChRs) composed of known receptor subunits have yielded conflicting results. The use of knockout mice that lack α5, β2, α5β2 or α5β2α7 nAChR subunits enabled us to measure the single-channel properties of distinct α3β4, α3β4α5 and α3β4β2 receptors in superior cervical ganglion (SCG) neurons. Using this approach, we found that α3β4 receptors had a principal conductance level of 32.6 ± 0.8 pS (mean ± SEM) and both higher and lower secondary conductance levels. α3β4α5 receptors had the same conductance as α3β4 receptors, but differed from α3β4 receptors by having an increased channel open time and increased burst duration. By contrast, α3β4β2 receptors differed from α3β4 and α3β4α5 receptors by having a significantly smaller conductance level (13.6 ± 0.5 pS). After dissecting the single-channel properties of these receptors using our knockout models, we then identified these properties – and hence the receptors themselves – in wild-type SCG neurons. This study is the first to identify the single-channel properties of distinct neuronal nicotinic receptors in their native environment.