Persistent Phosphorylation of Synaptic Proteins following Middle Cerebral Artery Occlusion

Transient cerebral ischemia following 1 to 2 hours of middle cerebral artery occlusion (MCAO) in the rat leads to infarction, which can be diminished by synaptic transmission modulators, implying aberrant cell signaling in the pathogenetic process. The authors report here changes in the levels of tyrosine phosphorylated proteins (PTyr) and calcium calmodulin kinase II (CaMKII) phosphorylation of Thr 286, in synaptosomal, particulate, and cytosolic fractions of different cortical areas following 1 or 2 hours of MCAO, or 2 hours of MCAO followed by 2 hours of reperfusion. At the end of 2-hour MCAO, PTyr, and in particular the pp180, indicative of NR2A/B subunit, increased in the synaptosomal fraction in less ischemic areas while it decreased in more severe ischemic regions. During reperfusion, phosphorylation increased at least 2-fold in all reperfused areas. During 2 hours of MCAO, the phosphorylation of CaMKII increased 8- to 10-fold in the synaptosomal fraction in all ischemic brain regions. During reperfusion, the phospho-CaMKII levels remained elevated by approximately 300% compared with the contralateral hemisphere (control). There was no increase in phospho-CaMKII in the cytosolic fraction at any time during or following ischemia in any of the brain regions examined. The authors conclude that both tyrosine kinase coupled pathways, as well as CaMKII-mediated cellular processes associated with synaptic activity, are strongly activated during and particularly following MCAO. These results support the hypothesis that aberrant cell signaling may contribute to ischemic cell death and dysfunction, and that selective modulators of cell signaling may be targets for pharmacological intervention against ischemic brain damage.

The effect of thyroxine isomers on the processes of free radical oxidation in the subcellular fractions of the rat cerebral cortex

Effective concentrations for D and L thyroxin isomers were determined by the chemiluminescent (CL) method and their effects on free radical oxidation in the mitochondrial and synaptosomal fraction of adult rat cerebral cortex were studied in vitro. A OA DT4 in a model system with riboflavin was 2.2 times higher than L-T4. Effective concentrations for both thyroxin forms were 1$q= = 7.43 x 10~5+/-0.71 M for D-T4 and 15q=15.47 x 10~5+/1.23 M for L-T4. Thyroxin effect on membranous fraction of the brain cortex was studied in vitro using luminol-dependent peroxide CL. In normal concentrations (1 x 108 M) both hormone forms exerted equally intensive antioxidant effect which was more pronounced in the mitochondrial fraction, where CL decreased by 69 and 66%, while in the synaptosomal fraction it decreased only by 45 and 46%. Since D form possesses no hormonal activity, this effect may be due to phenol origin of thyroxin.