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Fig. 3. Inhibition of endocytosis of PrP^C on sensory neurons by RAP and siRNA^LRP1. (A,B) 1 µM RAP (B; A is vehicle control) inhibits the endocytosis of neuronal surface-labelled PrP^C (red) but not Tf (green) after 2 minutes at 37°C. (C,D) Neurons treated for 90 minutes with 250 nM siRNA^Con (C) or siRNA^LRP1.1 (D) before endocytosing surface-labelled PrP^C (red) and Tf (green) for 2 minutes at 37°C. (E,F) Immunocytochemical labelling of surface PrP^C (red) and total LRP1 (green) in neurons preincubated for 90 minutes with 250 nM siRNA^Con (E) or siRNA^LRP1 (F), used to assess the effect of LRP1 knockdown; data in supplementary material Tables S2c and S2d. Bars, 5 µm. (G,H) Reduction of LRP1 protein shown by immunoblot (G; the 515-kDa band is shown) and quantitated in H (mean band intensity normalised to actin, ±s.d., n=4 independent knockdown experiments) after 250 nM penetratin-siRNA addition for the times shown (plus a 4-hour point showing the effect of adding additional 250 nM siRNA at 2 hours). The samples shown in G were from the 4-hour time point with two additions of siRNA.

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