Quantification of astrocytic synaptic pruning in hippocampus in response to in vitro Aβ oligomer incubation via colocalization analysis with C1q

Abstract

Quantification of synaptic engulfment is an indirect measurement of synaptic pruning. Here, we provide a detailed protocol for the volumetric rendering of individual high-resolution astrocytes in the CA1 region of hippocampus in an in vitro slice model of Amyloid-beta (A{beta}) treatment. This includes free floating slice preparation, treatment with A{beta} oligomers, immunofluorescence, confocal imaging and analysis of individual astrocytes. We also provide a comprehensive analysis for 3D rendering of astrocytes and assessment of synaptic engulfment via "eat-me tag" C1q protein and synaptic marker PSD95. HighlightsO_LIMeasurement of synaptic engulfment in response to treatment with A{beta} peptide C_LIO_LIVolumetric reconstruction of high resolution individual astrocyte C_LIO_LIColocalization analysis of astrocyte and complementary "eat-me" protein C1q C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=193 SRC="FIGDIR/small/491159v1_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@17d70b6org.highwire.dtl.DTLVardef@72db3dorg.highwire.dtl.DTLVardef@97968aorg.highwire.dtl.DTLVardef@1c447b_HPS_FORMAT_FIGEXP M_FIG C_FIG