FIGURE 8. 6-OHDA also causes CRE repression with increased cytoplasmic to nuclear CREB ratios in SH-SY5Y cells.

A human neuroblastoma-derived dopaminergic cell line was treated with LD₅₀ doses of 6-OHDA (150 μM) for 3 h. A, in some experiments, cells were transfected with CRE-luciferase reporter 24 h earlier, and lysates were analyzed in quadruplicate wells for luciferase activity. Data are expressed as -fold induction of luciferase activity normalized to the untreated control (UT) group ± S.D. B, equal amounts of protein were loaded from cytoplasmic and nuclear fractions of lysates from cells treated as indicated. Immunoblot analysis for pCREB, total CREB, and loading controls were performed. C, SH-SY5Y cells were treated with vehicle or 6-OHDA for 3 h and subjected to immunofluorescence for pCREB. Note that 6-OHDA-treated cells display increased cytoplasmic pCREB staining, which shows a clumped distribution in contrast to the diffuse light pCREB staining in control cells. D, 6-OHDA elicits increased caspase 3 activity in SH-SY5Y cells that is significantly attenuated by co-treatment with Bt₂cAMP. A and D: *, p < 0.05 versus untreated control; **, p < 0.05 versus 6-OHDA alone (ANOVA followed by Student’s t test with Bonferroni correction).