It is important to note that some studies showed no synaptic bouton loss (Phinney et al., 1999), normal neurite outgrowth (Harper et al., 1998) and a near two-fold increase in spine density in APP deficient neurons (Bittner et al., 2009). The reason for these inconsistent results is unclear but may be due to the different APP−/− mice lines, strain background, and types of neuron that were assessed, and the methods used for quantifying the spines (both in vitro and in vivo). Here, we complemented the studies in cultured hippocampal neurons from APP−/− mice with appropriate rescue experiments. Further, in both in vitro and in vivo experiments, we directly measured dendritic spines rather than using surrogate measurements, such as synaptophysin immunostaining. More specifically, we cultured hippocampal neurons from APP−/− mice instead of RNAi knockdown, which may not deplete APP completely, and scored actual spine numbers rather than immunostained for synaptic markers. In addition, we used mouse instead of human APP to generate sAPP to validate the importance of sAPPα on synaptic morphology. Furthermore, we did not observe such rescue effects on
