Fig. 4

Higher vulnerability of SN dopaminergic neurons from NCS-1 knockout mice in a Parkinson’s disease mouse model. a Left: Tyrosine hydroxylase (TH) immunostaining of coronal midbrain sections of NCS-1 wildtype and NCS-1 knockout mice, repeatedly treated with MPTP/probenecid or saline as controls, as indicated. Middle: Stereological quantification of SN dopaminergic and VTA dopaminergic neurons (NCS-1 wildtype saline: n = 14; NCS-1 wildtype MPTP: n = 13; NCS-1 knockout saline: n = 10; NCS-1 knockout MPTP: n = 11), and relative remaining neurons in MPTP-treated mice. Right: Mean absolute counted numbers of SN dopaminergic neurons in all analyzed sections for all animals, section position according to bregma. Error bars: SEM. b Left: TH immunostaining of striatal sections of NCS-1 wildtype and NCS-1 knockout mice, repeatedly treated with MPTP/probenecid or saline as controls, as indicated. Middle: Optical density quantification of TH signal in dorsal striatum (DS) and ventral striatum (VS) (NCS-1 wildtype saline: n = 14; NCS-1 wildtype MPTP: n = 13; NCS-1 knockout saline: n = 10; NCS-1 knockout MPTP: n = 11), and relative remaining TH signal in MPTP-treated mice. Right: Mean intensities for all analyzed sections for all animals, section position according to bregma. Error bars: SEM. c Left: Striatal injection site documentation of an in vivo retrogradely traced aged (1.5 years) mouse, aligned with the mouse brain atlas, and a labeled SN dopaminergic neuron, before UV-laser microdissection in fluorescent and brightfield mode. Scale bars: 10 µm. Right: Cell-specific relative mRNA levels of Cav2.3 in single SN dopaminergic neurons from juvenile (wildtype: n = 14; knockout: n = 20) and aged (1.5 years; wildtype: n = 15; knockout: n = 14) NCS-1 wildtype and NCS-1 knockout mice. d Cell-specific absolute UV-laser microdissection and reverse transcription quantitative PCR-based transcript molecule quantification in single mouse SN dopaminergic neurons for NCS-1 (juvenile: n = 13; adult: n = 14). e Left: Confocal images showing NCS-1 antibody staining (red) of TH-positive (green) neurons in SN and VTA of an adult wildtype mouse, respectively. Scale bar: 10 µm. Middle: Histogram showing the immunosignal intensity distributions of cytoplasmic NCS-1 signal, and respective background signal intensities for all analyzed TH positive SN (red) and VTA (blue) neurons, exemplary for one C57BL/6J mouse (SN: n = 157; VTA: n = 169). Right: Mean NCS-1 immunosignal quantification in SN and VTA dopaminergic neurons for all analyzed mice (n = 4). Antibody specificity was confirmed on NCS-1 knockout mice. Tukey's boxplots are shown. Significances are indicated by asterisks: *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Data values and additional bootstrapping analysis are detailed in Supplementary Tables 8C/D, 4D/E, 5A/B and Supplementary Figs. 4, 6. Source data are available as a Source Data file