Preferential localization of muscarinic M1 receptor on dendritic shaft and spine of cortical pyramidal cells and its anatomical evidence for volume transmission.

Acetylcholine (ACh) plays important roles for higher brain functions, including arousal, attention, and cognition. These effects are mediated largely by muscarinic acetylcholine receptors (mAChRs). However, it remains inconclusive whether the mode of ACh-mAChR signaling is synaptic, so-called "wired," transmission mediated by ACh released into the synaptic cleft, or nonsynaptic, so-called "volume," transmission by ambient ACh. To address this issue, we examined cellular and subcellular distribution of M(1), the most predominant mAChR subtype in the cerebral cortex and hippocampus, and pursued its anatomical relationship with cholinergic varicosities in these regions of adult mice. M(1) was highly expressed in glutamatergic pyramidal neurons, whereas it was low or undetectable in various GABAergic interneuron subtypes. M(1) was preferentially distributed on the extrasynaptic membrane of pyramidal cell dendrites and spines. Cholinergic varicosities often made direct contact to pyramidal cell dendrites and synapses. At such contact sites, however, synapse-like specialization was infrequent, and no particular accumulation was found at around contact sites for both M(1) and presynpatic active zone protein Bassoon. These features contrasted with those of the glutamatergic system, in which AMPA receptor GluA2 and metabotropic receptor mGluR5 were recruited to the synaptic or perisynaptic membrane, respectively, and Bassoon was highly accumulated in the presynaptic terminals. These results suggest that M(1) is so positioned to sense ambient ACh released from cholinergic varicosities at variable distances, and to enhance the synaptic efficacy and excitability of pyramidal cells. These molecular-anatomical arrangements will provide the evidence for volume transmission, at least in M(1)-mediated cortical cholinergic signaling.