Lead-Induced Dendritic Spine Growth Model

Since day DIV 7 to DIV 12 is the primary time period of dendritic spine growth (Zhao et al., 2018 (link)), therefore, to establish Pb-exposure model in vitro, 1 μM of Pb acetate (Sigma-Aldrich, United States) was added to the cultured hippocampal neurons for 5 days in this period. On DIV 12, the primary culture medium for the hippocampal neurons was replaced with sugar-free neurobasal medium and the neurons were further cultured for 2 h. Afterward, the neurons were cultured with 100 μM 2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl) amino)-6-deoxy-glucose (2-NBDG) and 1 μg/ml insulin for 30 min. The cells were subsequently collected for flow cytometry after being resuspended in PBS.