Extended Data Fig. 2: Further assessment of ECM and synapse dynamics in the developing hindbrain.
From: Extracellular matrix proteolysis maintains synapse plasticity during brain development
a) Representative images of brevican and hyaluronan (ubi:ssncan-GFP) in the hindbrain at 7, 14, 28, and 60 dpf. Scales: 100 µm. b) Representative image of PNN (perineuronal net)-like brevican staining at 60 dpf. Square indicates the inset on the right. Arrowheads indicate PNN-like brevican signals. Scale: 100 µm. c) Representative images of Chat-PSD95FingR puncta co-stained with presynaptic protein synapsin. Dashed circles indicate Chat-PSD95FingR puncta and arrowheads indicate colocalization with synapsin. Scale: 2 µm. d) Representative images of co-staining for brevican and Chat-PSD95FingR at 14 dpf. Dashed box indicates inset on the right. Arrowhead indicates association between brevican and Chat-PSD95FingR puncta. Dashed circle indicates location of Chat-PSD95FingR puncta. Scale: 5 µm. e) Representative images of co-staining for brevican and MBP (myelin basic protein) at 14 dpf hindbrain. Dashed line indicates the hindbrain region. Square indicates inset on the right. Scale: 50 µm. f) Representative images of Chat-PSD95FingR at 5, 9, and 13 dpf from the hindbrain of the same fish. Scales: 20 µm. g) Plots of lost (left, pink) and new (right, green) synapses for individual fish from Fig. 1j. Individual dots indicate the value for each fish (5-6 dpf, n = 15 fish; 7-8 dpf, n = 18 fish; 9-10 dpf, n = 18 fish; 11-12 dpf, n = 15 fish; 13-14 dpf, n = 17 fish). Asterisks in the figure represent results of Tukey’s multiple comparisons with respect to the 5-6 dpf shown in Fig. 1j. h) Quantification of percentage of new, lost, and stable synapses in each measured time point of Fig. 1j. Total number of synapses were calculated as the sum of the number of newly observed, lost and stable synapses. i) Quantification of synapse diameter from stable synapses and new synapses at t = 6 (Stable, n = 157 synapses from n = 14 fish; New, n = 25 synapses from n = 14 fish; p = 0.206, Welch’s t-test, performed for synapses). The synapse diameters were normalized by the mean value of stable synapses for each cell. Values were plotted as mean ± SEM. *: p < 0.05; **: p < 0.01; ns: not significant. For the representative images, similar results were observed in at least n = 2 experiments.
