Fig. 3. Overexpression of Synage in the cerebella of knockout mice rescues the numbers of both Purkinje cells and synapses to the level of wild-type mouse.

a Schematic representation of the AAV injection (AAV-EGFP-control and AAV-EF1α-Synage) and the analysis strategy for WT and KO mice. b, c Quantification of the number of Purkinje cells (PCs) per cerebellar sections (b) and representative immunofluorescence staining images of PCs (labeled with Calbindin, red) (c) in 3-week-old WT and KO mouse cerebella after stereotaxic injection of AAV-EGFP-control (control) or AAV-EF1α-Synage (Synage OE) into the neonatal mouse cerebella. Nuclei were stained with Hoechst 33342 (blue). The numbers (1 and 2) show the enlarged areas. Left scale bar: 200 μm; scale bar of the enlarged regions: 100 μm (WT Control, n = 7; WT Synage OE, n = 8; KO Control, n = 14; KO Synage OE, n = 11). d–e Representative transmission electron microscopy (TEM) micrographs of synapses upon AVV-mediated Synage overexpression (OE) on cerebellar cortex in adult WT and Synage KO mice. Synapses are indicated by red asterisks, upper scale bar: 500 nm (d), lower scale bar: 200 nm (e). f Quantification of the number of synapses per 15 μm² in three mice (WT Control, n = 93; WT Synage OE, n = 47; KO Control, n = 29; KO Synage OE, n = 53). g Quantification of the number of SVs per μm² in WT and KO mice (WT Control, n = 191; WT Synage OE, n = 84; KO Control, n = 70; KO Synage OE, n = 74). h–j Representative traces of mIPSCs from cerebellar PCs (h) and quantification of mIPSC amplitude (i) and frequency (j) in P26 WT and KO cerebella. Dots indicate data from individual experiments. k–m Representative traces of mEPSCs from cerebellar PCs (k) and quantification of mEPSC amplitude (l) and frequency (m) in P26 WT and KO cerebella. Dots indicate data from individual experiments.