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. 2020 Jun 18;5(12):e136007. doi: 10.1172/jci.insight.136007

Figure 4. LM rescued postischemic TrkB/PLCγ1 pathway activation, activated the TrkB/ERK1/2 pathway, and rescued ipsilateral KCC2 degradation.

Figure 4

All proteins of interest were normalized to housekeeping protein β-actin. Phosphoproteins were also normalized to their respective total protein (see Supplemental Figure 3). The * indicates differences between treatment group and naive controls; the # indicates differences between contralateral and ipsilateral hemispheres within groups. (A) Representative Western blots showing TrkB and p–TrkB-Y816 expression for all treatment groups. (B) Contralateral (L) and ipsilateral (R) TrkB expression 24 hours after ischemic insult. ***P < 0.001 by 1-way ANOVA. #P < 0.05 by 2-tailed t test. (C) Contralateral (L) and ipsilateral (R) p–TrkB-Y816 expression 24 hours after ischemic insult. **P < 0.01, ****P < 0.0001 by 1-way ANOVA. (D) Representative Western blots showing PLCγ1 and p–PLCγ1-Y783 expression. (E) Contralateral (L) and ipsilateral (R) PLCγ1 expression 24 hours after ischemic insult. #P < 0.05 by 2-tailed t test. (F) Contralateral (L) and ipsilateral (R) p–PLCγ1-Y783 expression 24 hours after ischemic insult. *P < 0.05, ***P < 0.001 by 1-way ANOVA. #P < 0.05 by 2-tailed t test. (G) Representative Western blots showing ERK1/2 and p-ERK1/2-T202/Y204 expression. (H) Contralateral (L) and ipsilateral (R) ERK1/2 expression 24 hours after ischemic insult. #P < 0.05 by 2-tailed t test. (I) Contralateral (L) and ipsilateral (R) p-ERK1/2-T202/Y204 expression 24 hours after ischemic insult. *P < 0.05, **P < 0.01 by 1-way ANOVA. (J) Representative Western blots showing KCC2 and p–KCC2-S940 expression. (K) Representative Western blot showing lack of KCC2 and p–KCC2-S940 band in liver samples from P7 pups. (L) Contralateral (L) and ipsilateral (R) KCC2 expression 24 hours after ischemic insult. #P < 0.05, ###P < 0.001 by 2-tailed t test. (M) Contralateral (L) and ipsilateral (R) p–KCC2-S940 expression 24 hours after ischemic insult. #P < 0.05, ##P < 0.01 by 2-tailed t test. Number of mice: n = 13 [6/7] (naive [M/F]), 19 [10/9] (ligate+PB), 13 [5/8] (post-LM), 22 [11/11] (pre-LM). Box-and-whisker plots show quartiles with median with minima and maxima at the bottom and top whiskers, respectively.