Fig. 1: Low concentrations of Aβo increased synapse density in secondary dendrites of primary cortical neurons.
(a) The strategy for revealing synaptic changes with expansion microscopy between the three experimental groups. At DIV 21, primary cortical neurons were daily treated for five days with Aβo (100 nM), or Aβo with eFT508 (2 nM); control cells were not treated. At DIV 26, the neurons were fixed and anchored, followed by embedding in a hydrogel matrix. The embedded cells were denatured at 60 °C, blocked, and labelled with specific antibodies for immunostaining. Confocal microscopy was used to image the expanded cells, and image analysis was performed. This figure panel was created using BioRender. (b) Representative images of control (Ctrl) neurons, and neurons treated with Aβo with or without eFT508. For immunostaining synapsin1 (Syn1, green) served as pre-synaptic marker, PSD95 (white) as post-synaptic marker, MAP2 (magenta) as marker of the neuronal structure. Scale bar: 70 µm. (c) Representative comparison of primary cortical neurons visualized before expansion (left), after expansion (middle), and in the 3D analysis model (right). For a chosen dendritic region single synapse bouton (SSBs), multi-innervated spines (MIS), multi-spine boutons (MSBs) including complex MSBs (cMSBs) containing more than 2 spines are indicated. Red arrow (upper left): SSB; yellow arrow (bottom left): MSB; green arrow (upper right): complex MSB; blue arrow (bottom right): MIS. Scale bar: 70 µm. (d) Aβo significantly increased synapse density compared to the control group and eFT508 suppressed this effect (F2, 33 = 7.513, p = 0.002; Ctrl vs. Aβo: q = 5.482, p = 0.0014). Data represent mean ± SEM, n = 6 biological replicates and analysis of 2 dendritic segments per biological replicate. **p < 0.01, ns = not significant.
