To investigate the effects of fibrillar Aβ(1-40) on the morphology and viability of cholinergic neurons and the involvement of the insulin-signaling pathway, we established primary cultures of rat basal forebrain cholinergic neurons and observed their responses to treatment with fibrillar Aβ(1-40) at different concentrations for different durations. Cell morphology was examined under microscope after immunofluorescence staining for neurofilament protein, cell vitality accessed by the Methyl thiazolyl tetrazolium assay, and expressions of a panel of insulin signaling-related proteins was detected by Western blot analysis. We show here that, at low concentrations of 0.1-1.0 micromol/L, fibrillar Aβ(1-40) had little effects on the cells; however, at higher concentrations of 2-10 μmicromol/L, it caused pathological changes, decreased the cell viability, and reduced the expression of insulin receptor, insulin receptor substrate-I, Protein Kinase B, and B cell lymphoma/leukemia-2 in a dose- and time-dependent manner. These results demonstrate that fibrillar Aβ(1-40) not only decreases the viability of cholinergic neuron but also down regulates the expression of important proteins in the insulin signal transduction pathway. We speculate that fibrillar Aβ(1-40) may contribute to the pathogenesis of Alzheimer's through disrupting the insulin signaling pathway, therefore decreasing neuronal activity and eventually leading to the apoptosis and cell loss.
2010 Wiley-Liss, Inc.