A single intrastriatal injection of synthetic α-synuclein fibrils (PFFs) in wildtype non-transgenic mice led to the transmission of pathological α-synuclein and Parkinson-like Lewy pathology in interconnected brain regions. This process resulted in progressive loss of dopamine neurons in the substantia nigra pars compacta (SNpc), reduced dopamine levels, and motor deficits, establishing a mechanistic link between α-synuclein pathology and Parkinson’s disease (PD) features. Pathological α-synuclein spread through cell-to-cell transmission along neural connections, leading to the accumulation of Lewy bodies and neurites in regions such as the striatum, neocortex, and olfactory bulb. The spread was time-dependent, with pathology initially confined to the injected hemisphere and later spreading to other regions. α-Synuclein pathology in the SNpc was associated with the gradual loss of TH-immunoreactive neurons and dopamine neuron loss, which preceded the loss of striatal dopamine levels. Despite the spread of α-synuclein pathology, DA neurons in the ventral tegmental area (VTA) and other regions remained unaffected, suggesting that connectivity determines susceptibility. Motor deficits, including impaired coordination and balance, were observed in PFF-injected mice, consistent with PD-like symptoms. These findings demonstrate that pathological α-synuclein can initiate a neurodegenerative cascade in non-transgenic mice, recapitulating key features of PD. The study also shows that α-synuclein pathology spreads independently of genetic background, highlighting its role in PD pathogenesis.A single intrastriatal injection of synthetic α-synuclein fibrils (PFFs) in wildtype non-transgenic mice led to the transmission of pathological α-synuclein and Parkinson-like Lewy pathology in interconnected brain regions. This process resulted in progressive loss of dopamine neurons in the substantia nigra pars compacta (SNpc), reduced dopamine levels, and motor deficits, establishing a mechanistic link between α-synuclein pathology and Parkinson’s disease (PD) features. Pathological α-synuclein spread through cell-to-cell transmission along neural connections, leading to the accumulation of Lewy bodies and neurites in regions such as the striatum, neocortex, and olfactory bulb. The spread was time-dependent, with pathology initially confined to the injected hemisphere and later spreading to other regions. α-Synuclein pathology in the SNpc was associated with the gradual loss of TH-immunoreactive neurons and dopamine neuron loss, which preceded the loss of striatal dopamine levels. Despite the spread of α-synuclein pathology, DA neurons in the ventral tegmental area (VTA) and other regions remained unaffected, suggesting that connectivity determines susceptibility. Motor deficits, including impaired coordination and balance, were observed in PFF-injected mice, consistent with PD-like symptoms. These findings demonstrate that pathological α-synuclein can initiate a neurodegenerative cascade in non-transgenic mice, recapitulating key features of PD. The study also shows that α-synuclein pathology spreads independently of genetic background, highlighting its role in PD pathogenesis.