Figure 7

Proteolytic activation of protein kinase Cδ (PKCδ) in the mouse substantia nigra during LPS-induced dopaminergic degeneration. C57BL/6 mice were stereotaxically injected with a single dose of lipopolysaccharide (LPS) (5 μg) or saline in the substantia nigra (SN) to elicit dopaminergic degeneration. Mice were sacrificed after 14 days. (A) Proteolytic cleavage of PKCδ in mouse substantia nigra tissue. Western blots were performed on nigral tissue lysates obtained from mice stereotaxically injected with either LPS or saline. Strong proteolytic cleavage of PKCδ is seen in LPS-injected mice and was essentially absent in the saline-injected animals. Blots were also probed for tyrosine hydroxylase and β-actin to confirm accurate dissection of the nigral tissue and equal protein loading respectively. (B) PKCδ cleaved band intensity was quantified by densitometric analysis after normalization to β-actin (n = 4 to 6 mice per group). (C) PKCδ kinase activity in mouse substantia nigra tissue. PKCδ IP-kinase assays were performed on nigral tissue lysates. Kinase activity was quantified by densitometric analysis of the ³²P-histone bands (n = 4 mice per group). A representative kinase gel is shown. Western blots of tyrosine hydroxylase and β-actin were performed on the same lysates to verify accurate dissection of the nigral tissue. Data represent the group mean ± SEM. ** (P <0.01) indicates a significant difference between saline and LPS-injected mice.