Fig. 1

Experimental conditions required for detection of endogenous-PSER129. Rapid perfusion fixation was required for detection of endogenous-PSER129. Mice were euthanized by CO₂ inhalation, and blood cleared by transcadial perfusion with PBS. Following a delay of 30 or 60 min, mice were then perfused with 4% PFA. A PSER129 staining across the neuroaxis. B High magnification images of PSER129 staining in the hippocampus (HIP) and the OB mitral cell layer (MCL). C Proteins extracted from sections across the neuroaxis were resolved by SDS-PAGE and blotted. Blots were probed for total protein (Revert protein stain, Licor), αsyn, and PSER129. Quantification PSER129 (D) and αsyn (E) per protein normalized to the mean of optimal perfusion control. n = 4. ****ANOVA, Dunnett’s post-hoc test. p < 0.0001. Scale bar = 25 µm. Anesthetics did not influence PSER129 abundance in the brain. Mice were euthanized by CO₂ inhalation, and then rapidly fixed by transcadial perfusion with PBS followed by 4% PFA. Ten minutes prior to euthanasia, some animals were exposed to xylazine (XYL, 10 mg/kg) or Ketamine (KET, 100 mg/kg). F PSER129 staining in OB sections. G Proteins extracted from sections across neuroaxis were resolved by SDS-PAGE and blotted. Blots were probed for total protein (G top panel), αsyn, and PSER129. Quantification of total αsyn (H), total PSER129 (I), ratio of PSER129 to αsyn (J), and (K) correlation between αsyn and PSER129 content. No significant differences were found between experimental groups (One-way ANOVA, Tukey’s post-hoc). Total asyn did not correlate with total PSER129 (Pearson Correlation, R² = 0.01045). All sections developed with nickel-DAB Chromogen (black/purple) and counterstained with methylgreen. n = 4–5