*2.5. Activation of VEGF*/*VEGFR2 Regulates the PI3K*/*Akt Survival Signaling Pathway in the Hippocampal Neurons Following Experimental Status Epilepticus*

The activation of VEGF/VEGFR2 can subsequently activate PI3K/AKT signaling by altering the activation state of numerous downstream proteins relevant to cell proliferation and inhibiting apoptosis [34,35]. To determinate whether the PGC-1α regulates the survival signaling pathway, PI3K/Akt, through VEGF/VEGFR2 activation in the hippocampus following KA-induced status epilepticus, we delineated the regulatory effects of the PGC-1α activator and siRNA against *pgc-1*α on PI3K/Akt signaling. In the experiment, the phosphorylation state of PI3K (p-PI3K) and AKT (p-AKT), and total levels of PIK3 and AKT were measured by western blotting analysis. The ratio of p-PI3K/total PI3K and p-AKT/total AKT were raised 6 h after microinjection of 3% dimethyl sulfoxide (DMSO) and KA into the left hippocampal CA3 region compared with controls (Figure 6a,b).

In addition, the ratio of p-PI3K/total PI3K and p-AKT/total AKT were significantly enhanced 6 h after microinjection of resveratrol (100 μmol) followed by KA administration into the left hippocampal CA3 region (Figure 6a,b). On the other hand, the ratio of p-PI3K/total PI3K and p-AKT/total AKT were raised 6 h after administration of control siRNA with KA into left hippocampal CA3 region compared with the control group (Figure 6c,d). However, the ratio of p-PI3K/total PI3K and p-AKT/total AKT were inhibited in the hippocampal CA3 neuronal cells 24 h before pre-treated microinjection of siRNA against *pgc-1*α (2 μg) followed with KA microinjection into the hippocampus. Akt can be activated by p-PI3K and active PI3K (Figure 6c,d).

**Figure 5.** (**a**) Changes of *vegfr2* mRNA expression and (**b**) changes of VEGFR2 protein in the hippocampal CA3 tissues 6 h after administration with kainic acid (KA; 0.5 nmol) with pre-treatment with resveratrol (Res; 100 μmol), or 3% DMSO into bilateral CA3 subfield. (**c**) The *vegfr2* mRNA expression in the hippocampus 6 h after the administration of KA into the left CA3 area 24 h after pre-treatment of control siRNA or *pgc-1*α siRNA (2 μg) into the bilateral CA3 subfield. (**d**) Changes in VEGFR2 protein expression in the CA3 tissue 6 h after the same treatments. Values are presented as mean ± SEM from four animals per experimental group. \* *p* < 0.05 versus control groups, + *p* < 0.05 versus DMSO+KA group, and # *p* < 0.05 versus control siRNA+KA group in the Scheffé multiple-range test.

**Figure 6.** (**a**) Changes of phosphorylated phosphatidylinositol 3-kinase (p-PI3K), total PI3K, phosphorylated AKT (p-AKT), and total AKT expression and (**b**) p-PI3K protein levels relative to total PI3K and p-AKT relative to total AKT in the CA3 region of the hippocampus 6 h after microinjection of kainic acid (KA; 0.5 nmol) with pretreatment of microinjection of 3% DMSO or resveratrol (Res; 100 μmol) into the hippocampal CA3 subfield. (**c**) Ratio of activated phosphorylation of PI3K and AKT levels in the CA3 subfield of the hippocampus 6 h after the application of KA (0.5 nmol) into the left side of the hippocampal CA3 region 24 h before treatment with control siRNA or siRNA for *pgc-1*α (2 μg) application into the bilateral CA3 subfield. (**d**) Phosphorylation levels of PI3K and AKT in the CA3 region of the hippocampus after 6 h of the same experiments. Values presented as mean ± SEM from six animals per experimental group. \* *p* < 0.05 versus the control group, + *p* < 0.05 versus DMSO+KA group, and # *p* < 0.05 versus control siRNA+KA group in the Scheffé multiple-range test.
