Fig. 13. Overexpression of NGF results in missorting of NGF from the constitutive to the regulated secretory pathway.a, AtT 20 cells were infected for 1 hr with 1 (A), 5 (B), or 10 (C) MOI of VV coding for pro-NGF, postinfected for 8 hr, fixed, and prepared for immunocytochemistry using an antibody to NGF followed by a CY3-conjugated secondary antibody. Cells were analyzed by confocal microscopy. b, AtT 20 cells were infected with 1 (A), 5 (B), or 10 (C) MOI for 1 hr followed by a 4 hr postinfection and 3 hr incubation in medium containing [35S] Translabel. Conditioned media were collected (CM1), the cells were chased for 3 hr, and media was again collected (CM2). 8-bromo-cAMP (5 mm) was then added to some cultures, and cells were incubated for an additional 3 hr, after which media were collected (CM3) and the cells were lysed (CL). NGF was immunoprecipitated from all samples, and the precipitate was analyzed by SDS-PAGE. Comparison of CM3 samples shows that NGF release can be stimulated by cAMP from cells infected with 5 or 10 MOI but not from cells receiving 1 MOI.