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Regulation of conventional protein kinase Cγ on synapsin-Ⅰ phosphorylation and their role in cerebral ischemic injury
ZHU Hong-yi ZHANG Nan YU Jin-ling WEI Hai-ping HAN Song LI Jun-fa*
Acta Anatomica Sinica ›› 2016 ›› Issue (2) : 145-151.
Regulation of conventional protein kinase Cγ on synapsin-Ⅰ phosphorylation and their role in cerebral ischemic injury
Objective To explore the regulatory effect of conventional protein kinase Cγ (cPKCγ) on synapsin-Ⅰ phosphorylation level and their role in oxygen-glucose deprivation (OGD) induced ischemic injury in primary cultured cortical neurons of mice. Methods By using the middle cerebral artery occlusion (MCAO) mouse model in vivo and OGD-induced ischemic model of primary cultured cortical neurons in vitro, we examined the interaction and regulation of cPKCγ on synapsin-Ⅰ phosphorylation, and the effect of cPKCγ on synaptic morphology after OGD injury through coimmunoprecipitation, Western blotting and immunofluorescence with the help of cPKCγ knockout (cPKCγ -/-) mice. Results cPKCγ interacted with synapsin-Ⅰ in both the intact and injured cerebral cortex of mice. Five possible serine phosphorylation cites were screened and results showed that only Ser549 and Ser553 were obviously changed after OGD and cPKCγ knockout. The statistic analysis further revealed that OGD insults significantly reduced phosphorylation of synapsin-Ⅰ at Ser549 and Ser553( n=5 per group，P<0.05, compared with cPKCγ +/+ normoxia group). The knockout of cPKCγ decreased synapsin-Ⅰ phosphorylation (n=5 per group, P<0.05, compared with cPKCγ +/+ normoxia group), while OGD further extended the decrease (n=5 per group，P<0.05, compared with cPKCγ +/+ OGD group). In addition, OGD treatment also decreased the length and number of neurites of primary cultured cortical neurons(n=8 per group, P<0.05, compared with cPKCγ +/+ normoxia group), and cPKCγ -/- group worsened the damage of synaptic morphology induced by OGD (n=8 per group, P<0.05, compared with cPKCγ +/+ OGD group). Conclusion cPKCγ may influence the morphology of neurite growth through regulating synapsin-Ⅰ phosphorylation at Ser549 and Ser553, thereby protect the cortical neurons against ischemic/hypoxic injury.
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