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Objective: To quantify genome-wide DNA methylation in the olfactory bulbs, fronto-parietal and occipital regions, and cerebellum in normal male Wistar rats and in modeling incomplete cerebral ischemia caused by permanent bilateral occlusion of the common carotid arteries., Material and Methods: The study was performed on 23 male Wistar rats divided into groups: «Sham operation» and «Cerebral ischemia». The level of genome-wide methylation of CCGG sites was determined by methyl-sensitive restriction using MspI/HpaII endonucleases followed by densitometric analysis of electrophoregrams by ImageJ software., Results: Incomplete cerebral ischemia on the 7th day leads to 56.3% (95% CI: 33.2-76.90) mortality. In the surviving rats of the «Cerebral ischemia» group, compared with the animals of the «Sham operation» group, a pronounced neurological deficit was observed, which was accompanied by changes in the level of whole-genome DNA methylation in the nervous tissue of brain structures ( p <0.05). Incomplete cerebral ischemia in male Wistar rats was characterized by interhemispheric asymmetry in the severity and direction of the epigenomic reaction of the nervous tissue in both ischemic and non-ischemic areas of the brain., Conclusion: It is likely that it is precisely this dynamics of changes in the status of genome-wide DNA methylation in the nervous tissue that imparts plasticity to neuronal function during ischemic damage.

The enrichment of angioneurology with fundamental advances leads to the understanding of new important facets in the pathogenesis of cerebral ischemia. The knowledge of epigenetic mechanisms in the development of stroke, in particular, DNA methylation, which makes a significant contribution to the development and formation of cerebral damage, is becoming more and more relevant. This review reflects an analysis of animal studies proving the relationship of DNA methylation with cerebral ischemia. As a result of the search work, 282 articles from the PubMed database were selected for keywords that corresponded to this topic. Of these publications, 8 studies were devoted to genome-wide DNA methylation, and 6 published the results of DNA methylation of candidate genes in experimental cerebral ischemia. The results have demonstrated that brain DNA methylation in animals is associated with the development of ischemic stroke and may play a role in several pathogenetic mechanisms. In two studies, a decrease in the level of DNA methylation in 2 genes in ischemic brain tissues of laboratory animals was found, at the same time, in four studies, 8 genes, in which methylation increased after ischemic stroke, were reported. These data suggest that the assessment of the level of DNA methylation in stroke is a promising biomarker for the search and improvement of pharmacological and non-pharmacological methods for limiting brain damage in ischemic and reperfusion injury at the stages of preclinical and clinical studies.

Bcl-2 protein expression was studied in hippocampal CA1, CA2, CA3 and CA4 pyramidal neurons in Mongolian gerbils (Meriones unguiculatus), of the in in the early (Day 2) and late (Day 7) reperfusion period after a 7-minute forebrain ischemia and following ischemic postconditioning (IPostC), as well as in sham-operated animals (n=60). In the latter, the highest level, of Bcl-2-expression was found in CA4 neurons, while the lowest--in-CA1 neurons (P<0.01). Reversible ischemic brain damage led to the increasing deficit of morphologically unchanged hippocampal neurons with the increasing duration of reperfusion period. This was accompanied by a significant decrease in expression of Bcl-2 in the early reperfusion period, but in the late reperfu- sion period this decrease largely disappeared. IPostC, applied as three episodes of ischemia-reperfusion lasting 15/15 seconds, contributed to significant increase in the number of morphologically unchanged CA1 and CA3 neurons in the early reperfusion period, while the expression of Bel-2 was increased in morphologically unchanged neurons in all the hippocampal areas. In the late reperfusion period after IPostC, the number of unchanged neurons was increased in hippocampal areas CA1, CA3 and CA4 (P<0.05), while a significant increase in Bcl-2 expression (by 12.7%, P<0.01) was detected only in CA1 neurons. The results suggest that the cytoprotective effect of IPostC in hippocampal CA1 area is realized through a mechanism leading to increased expression of Bcl-2 protein, i.e., by blocking apoptosis.

The aim of this study was to determine the effect of ischemic postconditioning (IP) on the viability of neurons in various hippocampal areas as well as on cytoplasmic activity of succinatedehydrogenase (SDH) in these cells in 30 male Mongolian gerbils (Meriones unguiculatus). Ischemic brain injury was induced by bilateral common carotid artery occlusion for 7 min. IP protocol comprised 3 cycles of 15 s of reperfusion/15 s of ischemia. After reperfusion for 48 h, the morphometric analysis was conducted, and SDH cytoplasmic activity was assessed using quantitative histochemistry in the pyramidal neurons of the hippocampal areas CA1, CA2, CA3, CA4. The experiment has demonstrated that 7-minute-long ischemia resulted in a significant decrease in the number of viable neurons in CA1 area (up to 24%) and in the CA3 (to 56%) of the hippocampus; besides, it lead to the elevation of SDH activity in the cytoplasm of the neurons in all the hippocampal areas as compared to that in sham-operated animals. The application of IP significantly increased the number of viable neurons in CA1 (up to 52.9%, P<0,01) and in CA3 areas of the hippocampus(up to 88%, P<0,05), and it was accompanied by reduction of SDH activity in surviving neurons in all the hippocampal areas.

The present study was aimed to investigate neuroprotective effects of early and late ischemic preconditioning in the acute phase of ischemic brain damage in rats. It was found that a single five-minute ischemic episode of early ischemic preconditioning did not lead to significant neuroprotective effects in comparison with control group, while three five-minute ischemic episode early ischemic preconditioning accompanied by a significant increase in neurological deficit and growing damage rate in CA1 hippocampus neurons. In contrast, later ischemic preconditioning in form of single five-minute episode 24 hours before ischemia modeling, provided a significant neuroprotective effect, manifesting reduced neurological deficit and maintaining viability of CA1 hippocampus neurons.

A new rat model of global cerebral ischemia-reperfusion was proposed via reversible occlusion of the major vessels originating from the aortic arch and supplying the brain. This technique can be used for the search and study of exogenous (pharmacological) and endogenous methods of brain protection from ischemia-reperfusion injury.

Cerebral ischemia results in severe derangements of energy metabolism in the nervous tissue including activation of glycolytic pathway. Activity of cytosolic lactate dehydrogenase (LDH) in the specific brain structures remains unclear. The recent study was aimed at investigation into the LDH activity in the cytoplasm of both hippocampal and cortical neurons in Mongolian gerbils (Meriones unguiculatus) at different durations of reperfusion after global ischemia. Analysis showed that the activity of LDH in pyramidal neurons of various hippocampal areas and neurons of II, III and V cortical layers after 7-minute forebrain ischemia depended on both localization of the neurons and duration ofreperfusion. In general, the changes in postischemic cytosolic LDH activity include significant decrease in the LDH activity 2 days after reperfusion with varying degree of recovery on day 7 of reperfusion.

In the first part of this review molecular mechanisms of ischemic tolerance emerging as a result ofpreconditioning of the brain are discussed. Data on inductors, sensors, transducers and effectors of early and delayed ischemic tolerance are presented.

The aim of this study was to determine the effect of ischemic postconditioning on hippocampal CA1 neuronal survival and cytoplasmic activity of lactate dehydrogenase (LDH) in the gerbil model of cerebral ischemia-reperfusion injury. Ischemia was induced by bilateral common carotid artery occlusion (for 7 min) in male Mongolian gerbils (Meriones unguiculatus). Ischemic postconditioning protocol comprised 3 cycles of 15 s reperfusion/15 s ischemia. After 48 h of reperfusion, CA1 neuronal death was detected by Nissl staining and the cytoplasmic LDH was demonstrated histochemically in CA1 area of the hippocampus with a quantitative cytophotometric assessment of the enzyme activity. The results have shown that 7 min ischemia resulted in a significant decrease in the number of viable neurons (up to 24%) in the CA1 area of hippocampus; in addition, it reduced the activity of LDH in these neurons (from 0.260 +/- 0.009 to 0.190 +/- 0.006 relative units). The application of ischemic postconditioning significantly increased the number of viable neurons (up to 52.9%, P < 0.01) in the CA1 area of hippocampus, and it was accompanied by an increase in the activity of LDH (0.240 +/- 0.008 relative units, P < 0.001).

In the 2nd part the authors describe in details the main aspects of protective effect of preconditioning of the brain: inhibition of programmed cell death, weakening of phenomenon of excitotoxicity, activation of endogenous antioxidant systems, anti-inflammatory effects, modulation of glial cell function, changes in regional blood flow and vascular reactivity. In addition, data analysis on the impact of preconditioning on brain neurogenesis, the state of the blood-brain barrier, ion homeostasis and metabolism of neurons is presented. Review emphasizes the role of microRNAs in mechanisms of ischemic tolerance of brain. Profound understanding of molecular mechanisms of increased tolerance of brain to ischemic and reperfusion injury requires the implementation of this phenomenon in clinical practice.