Defective MF terminal targeting in Lnx1-null mice. (A) ZnT3 and NeuN staining showed robust expanded IPB axon terminal field and more terminal-contacted CA3 pyramidal cells in Lnx1^−/− mice compared with WT littermates. Bars: 200 µm (top); 30 µm (bottom). PCL, pyramidal cell layer. n = 7 mice per group. (B) Representative images showing spines of CA3 neuron (green) contacting terminals of MF axon (red) in PW3 Lnx1^−/− mice and control littermates. The higher magnification deconvolved 3D-rendered views (right) are as shown from regions of white box (left arrowhead). The overlapped spines and ZnT3⁺ terminals are defined as ZnT3⁺ synapses. Arrows indicate terminals on shaft. Bars: 100 µm (left); 2.5 µm (right). (C) Schematic showing terminals on spine (Terminal-Spine) or shaft (Terminal-Shaft). (D and E) Quantification of ratio for terminals on spine or shaft and ZnT3⁺ synapses determined from 3D-rendered views. n = 14–16 neurons from three to four mice for per group. (F and G) Representative and percentage of four types of pre- and postsynaptic dynamics in coculture system. Red labels, presynaptic axons; green labels, postsynaptic dendritic branches or spines. Arrowheads indicate contacted boutons or spines. Dotted line marks the area of bouton. *, AI ≥ 0.2; ns, AI < 0.2. n = 13–17 neurons. (H) The areas of newborn boutons are smaller when cocultured with Lnx1^−/− neurons (n = 17 boutons) than with WT neurons (n = 23 boutons). (I) Quantification of the dynamic areas of refined boutons in coculture system. n = 33–47 boutons per group. Means ± SEM; two-way ANOVA with Tukey’s post hoc test (I) or Student’s t test (A, D, E, G, and H); *, P < 0.05; **, P < 0.01; ***, P < 0.001.