Emerging evidence suggests that Parkinson's disease (PD) may have its origin in the enteric nervous system (ENS), from where α-synuclein (αS) pathology spreads to the brain^1-4. Decades before the onset of motor symptoms, patients with PD suffer from constipation and present with circulating T cells responsive to αS, suggesting that peripheral immune responses initiated in the ENS may be involved in the early stages of PD^1,5-7. However, cellular mechanisms that trigger αS pathology in the ENS and its spread along the gut-brain axis remain elusive. Here we demonstrate that muscularis macrophages (ME-Macs), housekeepers of ENS integrity and intestinal homeostasis, modulate αS pathology and neurodegeneration in models of PD^8,9. ME-Macs contain misfolded αS, adopt a signature reflecting endolysosomal dysfunction and modulate the expansion of T cells that travel from the ENS to the brain through the dura mater as αS pathology progresses. Directed ME-Mac depletion leads to reduced αS pathology in the ENS and central nervous system, prevents T cell expansion and mitigates neurodegeneration and motor dysfunction, suggesting a role for ME-Macs as early cellular initiators of αS pathology along the gut-brain axis. Understanding these mechanisms could pave the way for early-stage biomarkers in PD.