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. 2022 Mar 4;8(9):eabj0112. doi: 10.1126/sciadv.abj0112

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Copyright © 2022 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).

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Fig. 4. — (A) Example CA1 tertiary dendrites from control and 90 min after OGD slices, immunostained for gephyrin, with or without GSK3β-IX and PD98059. Scale bar, 2 μm. (B) Spine quantification (**P < 0.01, ***P < 0.001). Total spine density (spines per micrometer): Control, 1.41 ± 0.06; Control + PD + G.-IX, 1.46 ± 0.07; OGD, 1.09 ± 0.06; OGD + PD + G.-IX, 1.15 ± 0.05. (C) Quantification of gephyrin cluster density (***P < 0.001). (D) Quantification of gephyrin cluster volume. Mean number (A.U.): Control, 1.00 ± 0.08; Control + PD + G.-IX, 1.12 ± 0.08; OGD, 0.44 ± 0.08; OGD + PD + G.-IX, 1.16 ± 0.09 (***P < 0.001). Mean volume (A.U.): Control, 1.00 ± 0.04; Control + PD + G.-IX, 0.91 ± 0.05; OGD, 0.67 ± 0.04; OGD + PD + G.-IX, 0.96 ± 0.07. (E) Example CA1 pyramidal neurons biolistically transfected with tdTomato and gephyrin. Scale bar, 10 μm. (F) CA1 tertiary dendrites from control and 90 min after OGD slices transfected with either eGFP-gephyrin_WT or eGFP-gephyrin_S268A/S270A. (G) Spine quantification (*P < 0.05, **P < 0.01). Total spine density (spines per micrometer): Control + gephyrin_WT-GFP, 1.05 ± 0.052; Control + eGFP-gephyrin_S268A/S270A, 1.05 ± 0.090; OGD, eGFP-gephyrin_WT 0.74 ± 0.056; OGD + eGFP-gephyrin_S268A/S270A, 0.69 ± 0.064. (H) Gephyrin cluster density (**P < 0.01). Density: Control + eGFP-gephyrin_WT, 0.38 ± 0.58; Control + eGFP-gephyrin_S268A/S270A, 0.47 ± 0.092; OGD + eGFP-gephyrin_WT, 0.11 ± 0.015; OGD+ eGFP-gephyrin_S268A/S270A, 0.44 ± 0.12. (I) Gephyrin cluster volume (**P < 0.01). Mean volume: Control + eGFP-gephyrin_WT, 0. 85 ± 0.012; Control + eGFP-gephyrin_S268A/S270A, 0.22 ± 0.043; OGD + eGFP-gephyrin_WT, 0.046 ± 0.01; OGD+ eGFP-gephyrin_S268A/S270A, 0.21 ± 0.049. Data are shown as means ± SD. Two-way ANOVA with Bonferroni multiple comparison test. n = 18 slices.