6-OHDA leads to activation of the DNA damage repair pathway. Primary cultures prepared from Puma +/+ and -/- mice were treated with 20 μM 6-OHDA. A) Cultures were fixed 30 minutes after treatment and stained for TH and PAR. B) Cultures treated for 15 minutes, 30 minutes, or 30 minutes with 5 mM NAC pretreatment were fixed and stained for TH and PAR. TH-positive and PAR-positive cells were counted and the percentage of TH-positive cells that were also PAR-positive was calculated. Data were analyzed by one-way ANOVA (***, p < 0.001) with Bonferroni post-tests to compare each treatment to untreated (15 m: **, p < 0.01; 30 m: ***, p < 0.001). C and D) Cell lysates were collected in RIPA buffer at the indicated times and proteins levels were assessed by western blotting for p-Atm, p-Atr, p-p53, p53 and actin. E) Western blots in D were quantitated with ImageQuant. Levels of p-p53 were normalized to total p53 levels. Data were analyzed by two-way ANOVA (treatment: *, p < 0.05; genotype: ns). F) Cultures treated for 1 hour with 6-OHDA were fixed and stained for TH and p-p53.