Advances in transgenic technology as well as in the genetics of Alzheimer disease (AD) have allowed the establishment of animal models that reproduce amyloid-beta plaques and neurofibrillary tangles, the main pathological hallmarks of AD. Among these models, 3xTg-AD mice harboring PS1 (M146V), APP (Swe) and tau (P301L) human transgenes provided the model that most closely mimics human AD features. Although cortical cultures from 3xTg-AD mice have been shown to present disturbances in intracellular [Ca(2+)] homeostasis, the development of AD pathology in vitro has not been previously evaluated. In the current work, we determined the temporal profile for amyloid precursor protein, amyloid-beta and tau expression in primary cortical cultures from 3xTg-AD mice. Immunocytochemistry and Western blot analysis showed an increased expression of these proteins as well as several phosphorylated tau isoforms with time in culture. Alterations in calcium homeostasis and cholinergic and glutamatergic responses were also observed early in vitro. Thus, 3x-TgAD cortical neurons in vitro provide an exceptional tool to investigate pharmacological approaches as well as the cellular basis for AD and related diseases.