Fig. 6: Methods workflow.

A ROIs were placed in whole-slide scans of FFPE striatal sections. We selected ROIs in ACC- and PFC-innervated areas of the putamen (asterisks). DAT+, VGLUT1+, and PSD95+ synaptic density were quantified within the putaminal ROIs. The middle panel shows 3D reconstructions of DAT+, VGLUT1+, and PSD95+ synapses positive for different α-synuclein proteoforms. The bottom panel illustrates the quantification of synaptic size and synaptic pairs (pre- and post-synapse next to each other). B Dopaminergic neuronal density (TH+) and LB density were quantified in SN. C LB density was assessed in ACC and PFC. D Synaptosomes were isolated from frozen putamen in a subset of donors. E Total, pSer129, and CTT122 α-synuclein levels were quantified in synaptosome fractions using AlphaLISA immunoassays. Figure partially created in BioRender. Van de berg (2025) https://BioRender.com/6sljtqz. α-syn α-synuclein, A coupled with acceptor bead, aa amino acids, ACC anterior cingulate cortex, CTT122 C-terminal truncated α-synuclein at amino acid 122, D coupled with donor bead, DAT dopamine transporter, GM grey matter, HB homogenization buffer, IHC immunohistochemistry, LB Lewy body, LH left hemisphere, PFC prefrontal cortex, PSD95 post-synaptic density 95, SN substantia nigra, pSer129 α-synuclein phosphorylated at Serine 129, Syn1 α-synuclein detected at amino acid 91–99, RH right hemisphere, Tyrosine hydroxylase TH, VGLUT1 vesicular glutamate transporter 1.