Objective To explore the modulation of protein kinase during ischemia/reperfusion brain injury.Methods Primary hippocampal cultures were prepared from day-18 SD rat embryos. Hippocampal neurons were dissociated by incubation in typsin and purified with arabinosylcytosin (Ara-c), which could inhibit the proliferation of neuroglia. The postischemia time was simplified to 30 min and 60min. Changes of PKC activity in plasma and membrane were assessed by phosphoryl transfer pieces and the expression of PKCα protein was measured by Western blot.Results The activities of cytosolic PKC of the control group, ischemia 30 min group and ischemia 60 min group were (6.24±0.27) pmol/(min·mg), (3.26±0.21) pmol/(min·mg) and (3.05±0.17) pmol/(min·mg)respectively, while the activities of membrane PKC were (2.63±0.13) pmol/(min·mg), (8.85±0.32) pmol/(min·mg)and(10.63±0.35) pmol/(min·mg)respectively. After ischemia/reperfusion brain injury , the activities of cytosolic PKC were (0.97±0.19) pmol/(min·mg)and (0.82±0.16) pmol/(min·mg)respectively, while the activities of membrane PKC were (12.38±0.39) pmol/(min·mg)and (12.66±0.99) pmol/(min·mg)respectively. Ischemia and reperfusion injury significantly increased the activity of membrane PKC and decreased that of cytosolic PKC. These changes became more significant with the extension of ischmia duration. Similar results were also observed in the expression of PKCα protein.Conclusions Ischemia reperfusion injury of rats’ hippocampal neurons results in translocational activation of PKC, especially PKCα, and the activation might damage the neurons by promoting calcium overload.
Key words
ischemia brain injury /
reperfusion injury /
protein kinase
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References
[1] Zipfel G J, Babcock D J, Lee J M , et al. Neuronal apoptosis after CNS injury: the roles of glutamate and calcium [J]. J Neurotrauma, 2000, 17(10): 857-869.
[2] Zhai D, Li S, Wang M, et al. Disruption of the GluR2/GAPDH complex protects against ischemia-induced neuronal damage [J]. Neurobiol Dis, 2013, 54(6): 392-403.
[3] Bao L, Li R H, Li M, et al. Autophagy-regulated AMPAR subunit upregulation in in vitro oxygen glucose deprivation/reoxygenation-induced hippocampal injury [J]. Brain Res, 2017, 1668(8): 65-71.
[4] Chater T E, Goda Y. The role of AMPA receptors in postsynaptic mechanisms of synaptic plasticity [J]. Front Cell Neurosci, 2014, 8(11): 401-401.
[5] Plaza-Zabala A, Flores A, Martin-Garcia E, et al. A role for hypocretin/orexin receptor-1 in cue-induced reinstatement of nicotine-seeking behavior [J]. Neuropsychopharmacology, 2013, 38(9): 1724-1736.
[6] Newton A C. Protein kinase C: structure, function, and regulation [J]. J Biol Chem, 1995, 270(48): 28495-28498.
[7] Bright R, Raval A P, Dembner J M, et al. Protein kinase C delta mediates cerebral reperfusion injury in vivo [J]. J Neurosci, 2004, 24(31): 6880-6888.
[8] Kopach O, Viatchenko-Karpinski V, Atianjoh F E, et al. PKCα is required for inflammation-induced trafficking of extrasynaptic AMPA receptors in tonically firing lamina II dorsal horn neurons during the maintenance of persistent inflammatory pain [J]. J Pain, 2013, 14(2): 182-192.
[9] Ishida K, Kotake Y, Sanoh S, et al. Lead-Induced ERK Activation Is Mediated by GluR2 Non-containing AMPA Receptor in Cortical Neurons [J]. Biol Pharm Bull, 2017, 40(3): 303-309.
[10] Weiss S, Dascal N. Molecular aspects of modulation of L-type calcium channels by protein kinase C [J]. Curr Mol Pharmacol, 2015, 8(1): 43-53.
[11] Wang J Q, Guo M L, Jin D Z, et al. Roles of subunit phosphorylation in regulating glutamate receptor function [J]. Eur J Pharmacol,2014, 728(4):183-187.
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