Fig 1: BDNF from PCs acts on TrkB on CFs during CF synapse elimination. a Representative traces of CF-EPSCs recorded in a mOrange-positive control PC associated with EGFP-positive control CFs (top, from a P21 mouse) and a mOrange-positive BDNF-KD PC associated with EGFP–positive TrkB-KD CFs (bottom, from a P34 mouse). Scale bars, 10 ms and 1 nA. b Frequency distribution histograms for the number of CFs innervating each PC for the following six groups of PCs recorded in mice during P21–P46. (1) F(-) control of BDNF-PC-KD: fluorescence-negative control PCs sampled in the same slices for sampling BDNF-KD PCs (closed columns, n = 39 PCs from 11 mice); (2) F(-) control of TrkB-CF-KD: fluorescence-negative control PCs that were not associated with TrkB-KD CFs (gray columns, n = 46 PCs from 11 mice); (3) F(+) control: mOrange-positive control PCs associated with EGFP–positive control CFs (open columns, n = 30 PCs from 4 mice); (4) BDNF-PC-KD: PCs with BDNF-KD sampled in the same slices for sampling F(-) control PCs (green columns, n = 49 PCs from 11 mice), (5) TrkB-CF-KD: PCs associated with TrkB-KD CFs (blue columns, n = 64 PCs from 11 mice); (6) BDNF-PC-KD/TrkB-CF-KD: BDNF-KD PCs associated with TrkB-KD CFs (deep blue columns, n = 31 PCs from 5 mice). P = 0.643 among the three control groups (Kruskal–Wallis test with Dunn’s post-hoc test). P = 0.838 among the three KD groups (Kruskal–Wallis test with Dunn’s post-hoc test). P = 0.047 between F(+) control and BDNF-PC-KD/TrkB-CF-KD (Mann–Whitney U-test). F(+), fluorescent protein-positive; F(-), fluorescent protein-negative *P < 0.05, ns indicates no significant difference among the three groups
Fig 2: CF synapse elimination in TrkB-PC-KO and p75NTR-PC-KO mice. a Representative traces of CF-EPSCs in a PC from a control (left, P21) and a TrkB-PC-KO (right, P26) mouse. Holding potential was -10 mV. Single traces recorded at different stimulus intensities were superimposed. b Frequency distribution histogram for the number of CFs innervating each PC in control (open columns, n = 67 PCs from 7 mice) and TrkB-PC-KO (filled columns, n = 71 PCs from 7 mice) mice during P21–P31 (P = 0.612, Mann–Whitney U-test). ns indicates no significant difference between the two groups. c, d Similar to a, b but for results from p75NTR-PC-KO and control mice during P21–P41. CF-EPSCs in c were recorded in cerebellar slices from a control (left, P39) and a p75NTR-PC-KO (right, P38) mouse. Sample sizes for d were 54 PCs from 6 control mice and 69 PCs from 8 p75NTR-PC-KO mice (P = 0.548, Mann–Whitney U-test). ns indicates no significant difference between the two groups. Scale bars: a and c, 10 ms and 1 nA
Fig 3: Effect of knockdown of TrkB or p75NTR in inferior olivary neurons on CF synapse elimination. a Schema of vector constructs for TrkB-KD, TrkB-Res and p75NTR-KD. b, c Bright field (BF) and fluorescent images showing the expression of EGFP in inferior olivary neurons b and CFs in the cerebellum c from a P21 mouse. The box in b is enlarged in the inset. Dotted lines in c indicate the PC layer. Scale bars: b, 200 mm; c, 50 µm; inset in b, 20 µm. d Representative traces of CF-EPSCs recorded from a control PC (left, from a P21 mouse), a PC associated with TrkB-KD CFs (middle, from a P26 mouse) and a PC associated with CFs expressing both TrkB-KD and TrkB-Res (right, from a P21 mouse). Single traces recorded at different stimulus intensities were superimposed. Holding potential was -10 mV. Scale bars, 10 ms and 1 nA. e Frequency distribution histogram for the number of CFs innervating each PC for control PCs (open columns, n = 46 PCs from 11 mice), PCs associated with TrkB-KD CFs (blue columns, n = 64 PCs from 11 mice) and PCs associated with CFs expressing both TrkB-KD and TrkB-Res (light blue columns, n = 23 PCs from 3 mice) during P21–P59. (P = 0.0303, control vs. TrkB-CF-KD; P = 0.026, TrkB-CF-KD vs. TrkB-CF-Res/TrkB-CF-KD; P = 0.509, control vs. TrkB-CF-Res/TrkB-CF-KD, Mann–Whitney U-test). Control PCs and PCs associated with TrkB-KD CFs were recorded in the same mice. *P < 0.05. f, g Similar to d, e but for results from PCs associated with p75NTR-KD CFs and control PCs during P22–P25. CF-EPSCs in f were recorded in cerebellar slices from a control PC (left, from a P21 mouse) and a PC associated with p75NTR-KD CFs (right, from a P21 mouse). Scale bars, 10 ms and 1 nA. Sample sizes for g were 32 control PCs and 34 PCs associated with p75NTR-KD CFs in the same 5 mice (P = 0.832, Mann–Whitney U-test). ns indicates no significant difference between the groups
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