Aβ1–42 reduces synapse number and inhibits neurite outgrowth in primary cortical and hippocampal neurons: A quantitative analysis

Synaptic loss represents one of the earliest signs of neuronal damage and is observed within both Alzheimer's disease patients and transgenic mouse models of the disease. We have developed a novel in vitro assay using high content screening technology to measure changes in a number of cell physiological parameters simultaneously within a neuronal population. Using Hoechst-33342 to label nuclei, βIII-tubulin as a neuron-specific marker, and synapsin-I as an indicator of pre-synaptic sites, we have designed software to interrogate triple-labelled images, counting only those synaptic puncta associated with tubulin-positive structures. Here we demonstrate that addition of amyloid beta peptide (Aβ1–42), to either primary hippocampal or cortical neurons for 4 days in vitro has deleterious effects upon synapse formation, neurite outgrowth and arborisation in a concentration-dependent manner. Control reverse peptide showed no effect over the same concentration range. The effects of Aβ1–42 were inhibited by D-KLVFFA, which contains residues 16–20 of Aβ that function as a self-recognition element during Aβ assembly and bind to the homologous region of Aβ and block its oligomerisation. These effects of Aβ1–42 on synapse number and neurite outgrowth are similar to those described within AD patient pathology and transgenic mouse models.