Oligomeric Aβ Neurotoxicity and Testosterone Neuroprotection

Oligomeric Aβ was prepared according to our previous report [31 (link)]. Aβ_1–42 peptide was dissolved in 1,1,1,3,3,3-hexafluoro-2-propanol (Sigma) and then it was dried overnight by air at room temperature. The peptide pellet was resuspended with anhydrous DMSO (Sigma) at 2 mM as final concentration. Aβ was then bath-sonicated for 30 min at room temperature. Aliquot Aβ peptide was stored at −80°C deep freezer before use. The working concentration of Aβ was 5 μM in all experiments. This concentration of Aβ could induce synaptic degeneration including the reduction of synaptic proteins but not apoptosis when applied to hippocampal neurons for 24 h [28 (link), 31 (link)].
To investigate the neurotoxicity of Aβ on primary cultures of hippocampal neurons, Aβ was diluted with NB medium and was added to the cell culture at DIV14 for 24 h. Neuroprotective effects of testosterone were investigated in pretreatment experiments. 10 nM of testosterone has been considered to be at physiological dose [32 (link)]. Previous reports showed that 10 nM testosterone elicited neuroprotective effects in primary neuronal cultures [33 (link), 34 (link)]. Based on these findings, hippocampal neurons were incubated with 10 nM of testosterone (Sigma) for 1 h and then cotreated with oligomeric Aβ for 24 h. The control group was treated with DMSO as vehicle.