To assess changes in the level and localization of the proapoptotic proteins Pink1, Parkin, and Cofilin in rat dorsal root ganglia 24 hours after axotomy, we used the method of immunohistochemistry. To form an experimental model of axotomy, we dissected the rat sciatic nerve. As a control, we examined the ganglia of the intact nerve on the other side of the animal. It was demonstrated that in dorsal root ganglia, the proteins Pink1, Parkin, and Cofilin are localized in the cytoplasm of neurons. Their average level in the cytoplasm was significantly higher than the level in the nuclei. 24 hours after transection of the sciatic nerve, the mean level of Pink1, Parkin, and Cofilin proteins in the cytoplasm of neurons in the axotomized ganglion increased compared to the control. The Pink1/Parkin protein system is responsible for mitochondrial quality control. An increase in the expression of Pink1 and Parkin indicates that axotomy entails severe disturbances in the functioning of the mitochondrial network of neurons. Cofilin regulates the remodeling of the actin cytoskeleton by depolymerizing fibrillar actin during shape changes, cell movement, and endocytosis. Increased expression of Cofilin indicates that disruption of axon integrity triggers disassembly of actin filaments in the soma of neurons of dorsal root ganglia.
Ischemic tolerance determines resistance to lethal ischemia gained by a prior sublethal stimulus (i.e., preconditioning). We reproduced this effect in two variants. In vitro the preliminary short (5 s) photodynamic treatment (PDT) (photosensitizer Photosens, 10 nM, 30 min preincubation; laser: 670 nm, 100 mW/cm2) significantly reduced the necrosis of neurons and glial cells in the isolated crayfish stretch receptor, which was caused by following 30-min PDT by 66% and 46%, respectively. In vivo PDT of the rat cerebral cortex with hydrophilic photosensitizer Rose Bengal (i.v. administration, laser irradiation: 532 nm, 60 mW/cm2, 3 mm beam diameter, 30 min) caused occlusion of small brain vessels and local photothrombotic infarct (PTI). It is a model of ischemic stroke. Cerebral tissue edema and global necrosis of neurons and glial cells occurred in the infarction core, which was surrounded by a 1.5 mm transition zone, penumbra. The maximal pericellular edema, hypo- and hyperchromia of neurons were observed in penumbra 24 h after PTI. The repeated laser irradiation of the contralateral cerebral cortex also caused PTI but lesser as compared with single PDT. Preliminary unilateral PTI provided ischemic tolerance: at 14 day after second exposure the PTI volume significantly decreased by 24% than in the case of a single exposure. Sensorimotor deficits in PDT-treated rats was registered using the behavioral tests. The preliminary PTI caused the preconditioning effect.
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