Activation of Muscarinic Receptors Mediated Varying Cholinergic Effects on Medullar 5-HT Neurons and Locomotor Activities in ePet-EYFP Mice

Serotonergic neurons (5-HT) in medulla play an essential role in initiating locomotion. These neurons receive cholinergic input and display varying response to acetylcholine (ACh). However, the mechanism behind remains unknown. Using ePet-EYFP transgenic mice (P3–P6) and multiple approaches, we investigated mechanisms underlying cholinergic modulation of medullar 5-HT neurons and generating locomotion. Our results included: (1) Significant morphological difference in 5-HT neurons was shown between parapyramidal region (PPR) and midline raphe nuclei (MRN). (2) ACh induced triphasic effects (excitatory, inhibitory, and neutral) on 5-HT neurons, which could be duplicated by muscarine, with PPR neurons more excitable and MRN neurons less sensitive to ACh. (3) ACh enhanced excitability via mAChR-M3 receptors, while M2&M4 receptors mediated inhibitory effects. (4) Blocking M3 pathway reduced gait frequency and disrupted locomotion, whereas antagonizing M2&M4 pathways increased gait frequency. This study unveiled interactions between medullary cholinergic and serotonergic systems in modulating neuronal excitability and generating locomotion.