1 2 3 4 5 Cholinergic Modulation of Neuronal Excitability in the Rat Suprachiasmatic Nucleus 6 7 8 9

43 44 The central cholinergic system regulates both the circadian clock and sleep-wake cycle, and may 45 participate in the feedback control of vigilance states on neural excitability in the 46 suprachiasmatic nucleus (SCN) that houses the circadian clock. Here we investigate the 47 mechanisms for cholinergic modulation of SCN neuron excitability. Cell-attached recordings 48 indicate that the nonspecific cholinergic agonist carbachol (CCh) inhibited 55% and excited 49 21% SCN neurons, leaving 24% nonresponsive. Similar response proportions were produced by 50 two muscarinic receptor (mAChR) agonists, muscarine and McN-A-343 (M1/4 agonist), but not 51 by two nicotinic receptor (nAChR) agonists, nicotine and choline (α7-nAChR agonist), which 52 however produced similar response proportions. Whole-cell and perforated patch recordings 53 indicate that CCh inhibition of firing was mediated by membrane hyperpolarization due to 54 activation of background K currents, which were sensitive to submillimolar concentrations of 55 Ba and to millimolar concentrations of TEA. RT-PCR analysis demonstrated the presence of 56 mRNA for M1 to M5 mAChRs in SCN. The CCh-induced hyperpolarization and activation of 57 background K currents were blocked by M4 antagonists and to a lesser degree by M1 58 antagonists, but were insensitive to the antagonists for M2 or M3, suggesting the involvement of 59 M4 and M1 mAChRs in mediating CCh inhibition of firing. CCh enhancement of firing was 60 mediated by membrane depolarization, as a result of postsynaptic inhibition of background K 61 currents. The multiple actions of cholinergic modulation via multiple receptors and ion channels 62 may allow acetylcholine to finely control SCN neuron excitability in different physiological 63