42 (Bachem) oligomers were prepared as previously described (17). Aβ was resuspended in 1× Hanks’ balanced salt solution and DMSO and then placed in 4°C overnight. At DIV 14, primary hippocampal neurons were treated with 500 nM Aβ42 for 6 hours. Fasudil (Selleckchem, catalog no. S1573) and SR7826 (Tocris, catalog no. 562610) were reconstituted to a 10 mM stock in either water or DMSO, respectively. At DIV 14, primary hippocampal neurons were dosed with 10 μM SR7826, 30 μM fasudil, or a combination of drug and Aβ42 for 6 hours. Six hours was chosen on the basis of past studies demonstrating that Aβ42-induced spine loss in cultured neurons plateaus at about 6 hours after exposure (41), and pan-ROCK inhibitors induce robust changes in spine morphology on cultured hippocampal neurons after 6 hours of exposure (25). Blebbistatin (Tocris, catalog no. 1852) was reconstituted to a 10 mM stock in DMSO. At DIV 14, primary hippocampal neurons were treated with 5 μM blebbistatin for 1 hour. One-hour incubation time was selected on the basis of previous studies (72, 73).

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