Figure 5

Expression of Tau ^RDΔ causes morphological changes of mossy fiber boutons in hippocampal slice cultures. Photomicrographs of slice cultures from control (Ctrl) or pro-aggregant-mice (Pro) at DIV5 (a1-b3) and DIV10 (c1-d9); area CA3-immunostaining against pan-Tau (K9JA, a1,b1,c1,d1), phospho-Tau (12E8, a2,b2,c2,d2) and merged (a3,b3,c3,d3) pictures are depicted. (a1) No 12E8-immunoreactivity is present in control slices at DIV5 (a1-a3) and DIV10 (c1-c3). (b1) In contrast, in pro-aggregant slices 12E8-immunoreactivity is observed in pyramidal cell somata and dendrites at DIV5 (b1-b3) and DIV10 (d1-d3). (c4-c6) Zoom-in images (from (c1)) showing mossy fiber boutons filled with Tau. No 12E8 phosphorylation was detected in control slices. (d4-d6, zoom-in from d1) In contrast, boutons in pro-aggregant slices contain phosphorylated Tau. (c7-c9 + d7-d9) Immunostaining against pan-Tau (green), synaptophysin (S’physin, red) and merged picture is shown in control (c7-c9) and pro-aggregant slices (d7-d9). Arrows mark the axon shaft; boutons are marked by dotted line. (e) Higher magnification of single mossy fiber boutons (stained with DiI) in slices from control mice, pro-aggregant mice and pro-aggregant slices treated with compound bb14. (f) The bouton diameter was increased by ~42% in pro-aggregant slices in comparison to control and by only ~18% when compared to bb14-treated pro-aggregant slices. (g) The bouton surface was larger in pro-aggregant slices (by ~75%) vs. control and by only 32% vs. bb14-treated pro-aggregant slice cultures. (h) The average distance between boutons was increased by ~50% in pro-aggregant slices compared to control and bb14-treated pro-aggregant cultures. (i) Numbers of filopodia (control: 1.56; pro-aggregant: 1.89; pro-aggregant + bb14: 1.77) per bouton was only slightly affected in pro-aggregant slices. (j) The length of filopodia (control: 1.66 μm; pro-aggregant: 1.39 μm; pro-aggregant + bb14: 1.69 μm) was not changed in pro-aggregant slices. One way ANOVA followed by Tukey’s post-hoc test *** p- value < 0.001; * p-value < 0.05). Error bars represent SEM.