Fig. 4

Effect of human α-syn overexpression on glucose-derived carbon incorporation and turnover in the nuclear versus cytoplasmic compartment. a Representative SEM image of a large neuronal cell body located in the SNpc. b Segmentation of the SEM image into cytoplasmic (Cy), nuclear (Nu), and nucleolar (n) compartments. Outlines of the compartments were drawn manually using the Look@NanoSIMS software (see Additional file 1). c The mask generated in (b) was used to measure ¹³C enrichment in the nuclear versus cytoplasmic compartments in the corresponding isotopic NanoSIMS maps. Note that neurons were excluded from the analysis when the highly ¹³C enriched nucleolar compartment was present in the image. d The schematic illustrates the determination of the difference in ¹³C enrichment between the nuclear and cytoplasmic compartments. The graph compares neuronal cell bodies located in the control and α-syn-overexpressing SNpc at the 24 h, 48 h (pulse) and 96 h (chase) time points. Note the loss of nuclear glucose-derived ¹³C enrichment in the α-syn overexpression condition during the pulse period. The thick lines in the violin plots represent the median and the thin lines the upper and lower quartiles. The number of neurons analyzed is indicated at the bottom of the histogram bar. 24 h: N = 3 animals per group; 48 h: N = 4 animals per group; 96 h: N = 3 animals per group. Statistical analyses: multiple unpaired two-tailed t-test with False Discovery Rate (FDR) approach; FDR-adjusted P values are indicated in the graph. Scale bar: 5 µm