1 2 3 4 M 1 and M 4 receptors modulate hippocampal pyramidal neurons 5

31 Acetylcholine (ACh), acting at muscarinic ACh receptors (mAChRs), modulates the 32 excitability and synaptic connectivity of hippocampal pyramidal neurons. CA1 pyramidal 33 neurons respond to transient (" phasic ") mAChR activation with biphasic responses in 34 which inhibition is followed by excitation, whereas prolonged (" tonic ") mAChR activation 35 increases CA1 neuron excitability. Both phasic and tonic mAChR activation excites 36 pyramidal neurons in the CA3 region, yet ACh suppresses glutamate release at the 37 CA3-to-CA1 synapse (the Schaffer-collateral pathway). Using mice genetically lacking 38 specific mAChRs (mAChaR knockout mice), we identified the mAChR subtypes 39 responsible for cholinergic modulation of hippocampal pyramidal neuron excitability and 40 synaptic transmission. Knockout of M1 receptors significantly reduced, or eliminated, 41 most phasic and tonic cholinergic responses in CA1 and CA3 pyramidal neurons. On 42 the other hand, in the absence of other G q-linked mAChRs (M3 and M5), M1 receptors 43 proved sufficient for all postsynaptic cholinergic effects on CA1 and CA3 pyramidal 44 neuron excitability. M3 receptors were able to participate in tonic depolarization of CA1 45 neurons, but otherwise contributed little to cholinergic responses. At the Schaffer-46 collateral synapse, bath application of the cholinergic agonist carbachol suppressed 47 stratum radiatum-evoked excitatory postsynaptic potentials (EPSPs) in wild-type CA1 48 neurons and in CA1 neurons from mice lacking M1 or M2 receptors. However, 49 Schaffer-collateral EPSPs were not significantly suppressed by carbachol in neurons 50 lacking M4 receptors. We therefore conclude that M1 and M4 receptors are the major 51 mAChR subtypes responsible for direct cholinergic modulation of the excitatory 52 hippocampal circuit. 53 54 55 56