Figure 6.

shRNA knockdown of PAC1 hop expression. (a) pEAK rapid HEK-293 cells were transfected with control vector, or full length rat hop or short expression vectors without and with hop shRNAs. PAC1 antibody detects both hop and short proteins including both human as well as rat. Data were derived from three experiments. Transfection efficiency was 50–60% per group. (b) E10.5 precursors were transfected with either control shRNA or both hop shRNAs 2 h after plating, in the absence of PACAP (Vehicle) or the presence of PACAP (10 nM). BrdU was added at 22 h, and cells were fixed at 24 h. Cells that were transfected with control shRNA or hop shRNAs can be visualized by red fluorescence. The four groups were analyzed by one-way ANOVA, followed by Tukey's multiple comparison test. Six transfections from three experiments were performed, and transfection efficiency was 10–15% in E10.5 precursors. The number of cells analyzed from each experiment was 20–35 per dish. Scale bar, 10 μm. *p<0.05, **P<0.01.