title: Protein kinase C activity, translocation, and selective isoform subcellular redistribution in the rat cerebral cortex after in vitro ischemia.
abstract: Protein kinase C (PKC) involvement in ischemia-induced neuronal damage has been investigated in superfused rat cerebral cortex slices submitted to 15 min of oxygen-glucose deprivation (OGD) and in primary cultures of rat cortical neurons exposed to 100 microM glutamate (GLU) for 10 min. OGD significantly increased the total PKC activity in the slices, mostly translocated in the particulate fraction. After 1 hr of reperfusion, the total PKC activity was reduced and the translocated fraction dropped by 84% with respect to the control. Western blot analysis of OGD samples showed an increase in total beta(2) and epsilon PKC isoform levels. After reperfusion, the total levels of alpha, beta(1), beta(2) and gamma isoforms were significantly reduced, whereas the epsilon isoform remained at an increased level. Endogenous GLU release from OGD slices increased to about 15 times the basal values after 15 min of oxygen-glucose deprivation, and to 25 and 35 times the basal level in the presence of the PKC inhibitors staurosporine (0.1 microM) and bisindolylmaleimide (1 microM), respectively. Western blot analysis of GLU-treated cortical neurons showed a significant decrease only in the total level of beta(2) isoforms. Cell survival was reduced to 31% in GLU-treated neuronal cultures; PKC inhibitors were not able to modify this effect. These findings demonstrate that the cell response to OGD and GLU involves PKC in a complex way. The net role played by PKC during OGD may be to reduce GLU release and, consequently, neurotoxicity. The isoforms beta(2) and epsilon are affected the most and may play a significant role in the mechanisms underlying neurotoxicity/neuroprotection.
authors.affiliation: Department of Experimental and Diagnostic Medicine, Section of Medical Genetics, University of Ferrara, Italy. svr@dns.unife.it
authors.forename: Rita
authors.initials: R
authors.lastname: Selvatici
authors.forename: Silvia
authors.initials: S
authors.lastname: Marino
authors.forename: Carmela
authors.initials: C
authors.lastname: Piubello
authors.forename: Donata
authors.initials: D
authors.lastname: Rodi
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Beani
authors.forename: Enrico
authors.initials: E
authors.lastname: Gandini
authors.forename: Anna
authors.initials: A
authors.lastname: Siniscalchi
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D015153:Blotting, Western; D002545:Brain Ischemia / Q000201:enzymology*; D002470:Cell Survival / Q000502:physiology; D002540:Cerebral Cortex / Q000201:enzymology*; D046508:Culture Techniques; D018698:Glutamic Acid / Q000032:analysis; D007527:Isoenzymes / Q000378:metabolism*; D008297:Male; D011493:Protein Kinase C / Q000145:classification / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013347:Subcellular Fractions / Q000201:enzymology; D013997:Time Factors
chemical_list: D007527:Isoenzymes; D018698:Glutamic Acid; D011493:Protein Kinase C