Fig. 7: Aβ42o induces AMPK-dependent MFF phosphorylation at two serine sites required for Aβ42o-dependent dendritic mitochondrial fragmentation and spine loss.
a Representative images of secondary dendritic segments of primary cortical PNs at 21 DIV. Embryos at E15.5 were ex utero electroporated with pCAG-Venus, pCAG-mito-DsRed, MFF shRNA, and low levels of either pCAG-MFF-WT cDNA or phospho-dead pCAG-MFF-AA. At 20DIV, the neurons were treated with either a vehicle control or Aβ42o for 24 h. Gene replacement with a form of MFF that cannot be phosphorylated by AMPK (MFF shRNA + MFF-AA) blocks dendritic mitochondrial fragmentation and subsequent spine loss. b–d Quantification of b dendritic mitochondrial length, c dendritic mitochondrial density, and d spine density. All of the analyses were done blind to the experimental conditions and performed by manual counting using FIJI. Data are represented by box plots displaying minimum to maximum values, with the box denoting 25th, 50th (median), and 75th percentiles from three independent experiments. nMFFshRNA + MFFWT Control = 31 dendrites, 166 mitochondria; nMFFshRNA + MFFWT Aβ42o = 29 dendrites, 228 mitochondria; nMFFshRNA + MFFAA Control = 35 dendrites, 164 mitochondria; nMFFshRNA + MFFAA Aβ42o = 28 dendrites, 137 mitochondria. Statistical analyses were performed using a Mann–Whitney test in b–d. Exact P values are indicated on the figure when available through Prism software, otherwise, the test significance is provided using the following criteria: ****P < 0.0001. Scale bar in a: 2 μm.
