Nicotinic Transmission onto Layer 6 Cortical Neurons Relies on Synaptic Activation of Non-α7 Receptors.

Nicotinic excitation in neocortex is mediated by low-affinity α7 receptors and by high-affinity α4β2 receptors. There is evidence that α7 receptors are synaptic, but it is unclear whether high-affinity receptors are activated by volume transmission or synaptic transmission. To address this issue, we characterized responses of excitatory layer 6 (L6) neurons to optogenetic release of acetylcholine (ACh) in cortical slices. L6 responses consisted in a slowly decaying α4β2 current and were devoid of α7 component. Evidence that these responses were mediated by synapses was 4-fold. 1) Channelrhodopsin-positive cholinergic varicosities made close appositions onto responsive neurons. 2) Inhibition of ACh degradation failed to alter onset kinetics and amplitude of currents. 3) Quasi-saturation of α4β2 receptors occurred upon ACh release. 4) Response kinetics were unchanged in low release probability conditions. Train stimulations increased amplitude and decay time of responses and these effects appeared to involve recruitment of extrasynaptic receptors. Finally, we found that the α5 subunit, known to be associated with α4β2 in L6, regulates short-term plasticity at L6 synapses. Our results are consistent with previous anatomical observations of widespread cholinergic synapses and suggest that a significant proportion of these small synapses operate via high-affinity nicotinic receptors.