Amyloidogenic processing of the amyloid precursor protein (APP) by β- and γ-secretases generates several biologically active products, including amyloid-β (Aβ) and the APP intracellular domain (AICD). AICD regulates transcription of several neuronal genes, especially the Aβ-degrading enzyme, neprilysin (NEP). APP exists in several alternatively spliced isoforms, APP(695), APP(751), and APP(770). We have examined whether each isoform can contribute to AICD generation and hence up-regulation of NEP expression. Using SH-SY5Y neuronal cells stably expressing each of the APP isoforms, we observed that only APP(695) up-regulated nuclear AICD levels (9-fold) and NEP expression (6-fold). Increased NEP expression was abolished by a β- or γ-secretase inhibitor but not an α-secretase inhibitor. This correlated with a marked increase in both Aβ(1-40) and Aβ(1-42) in APP(695) cells as compared with APP(751) or APP(770) cells. Similar phenomena were observed in Neuro2a but not HEK293 cells. SH-SY5Y cells expressing the Swedish mutant of APP(695) also showed an increase in Aβ levels and NEP expression as compared with wild-type APP(695) cells. Chromatin immunoprecipitation revealed that AICD was associated with the NEP promoter in APP(695), Neuro2a, and APP(Swe) cells but not APP(751) nor APP(770) cells where AICD was replaced by histone deacetylase 1 (HDAC1). AICD occupancy of the NEP promoter was replaced by HDAC1 after treatment of the APP(695) cells with a β- but not an α-secretase inhibitor. The increased AICD and NEP levels were significantly reduced in cholesterol-depleted APP(695) cells. In conclusion, Aβ and functional AICD appear to be preferentially synthesized through β-secretase action on APP(695).