K. A. Baranova

Preconditioning induces prolonged expression of transcription factors pCREB and NF‐κB in the neocortex of rats before and following severe hypobaric hypoxia

Preconditioning using mild repetitive hypobaric hypoxia is known to increase a tolerance of brain neurons to severe hypoxia and other injurious exposures. In the present study, the effects of mild hypoxic preconditioning on the expression of transcription factors NF‐κB and phosphorylated CREB (pCREB) has been studied in the neocortex of rats exposed to severe hypobaric hypoxia. As revealed by quantitative immunocytochemistry, the injurious severe hypobaric hypoxia (180 Torr, 3 h) remarkably reduced the neocortical levels of pCREB and NF‐κB. The three‐trial hypoxic preconditioning (360 Torr, 2 h, 3 days) induced persistent up‐regulation of pCREB and NF‐κB expression in the neocortex of rats 3–24 h following the severe hypoxia. In addition, the preconditioning alone which was not followed by the severe hypoxia, considerably increased neocortical pCREB and NF‐κB levels. The findings suggest a role for transcription factors cAMP response element‐binding protein and NF‐κB in the neuroprotective mechanisms activated by the hypoxic preconditioning.

Mild hypobaric hypoxia preconditioning up-regulates expression of transcription factors c-Fos and NGFI-A in rat neocortex and hippocampus

Transcription factors c-Fos and NGFI-A encoded by immediate early genes largely participate in the biochemical cascade leading to genomically driven lasting adaptation by neurons to injurious exposures including hypoxia/ischemia. Present study was designed to examine the involvement of c-Fos and NGFI-A in the development of brain hypoxic tolerance induced by mild hypoxic preconditioning. Earlier we have reported that preconditioning by repetitive mild hypobaric hypoxia (MHH) considerably increases neuronal resistance to subsequent severe injurious exposures. Herein, changes of c-Fos and NGFI-A expression in vulnerable rat brain areas (hippocampus, neocortex) in response to preconditioning MHH itself were studied using quantitative immunocytochemistry. Exposure to MHH differentially enhanced c-Fos and NGFI-A expression in neocortex and hippocampal fields 3-24h following the last MHH trial. The c-Fos up-regulation was the most pronounced in neocortex, CA1, and dentate gyrus, but it was twice lower in CA3/CA4. The up-regulation of NGFI-A in CA1, dentate gyrus and neocortex was 1.5-2-fold lower than that of c-Fos; but in CA3 and CA4 the rates of the c-Fos and NGFI-A induction were comparable. The present findings indicate that cooperative but differential activation of c-Fos and NGFI-A expression in vulnerable brain areas contribute to the development of tolerance achieved by MHH preconditioning.

Mild hypobaric hypoxic postconditioning increases the expression of HIF-1α and erythropoietin in the CA1 field of the hippocampus of rats that survive after severe hypoxia

Using immunohistochemistry, we studied the expression of the alpha regulatory subunit of the hypoxia-inducible factor (HIF-1α) and the product of its target gene, which encodes the protective cytokine erythropoietin in the hippocampal CA1 field of rats in response to damaging severe hypoxia and severe hypoxia followed by three sessions of postconditioning with mild hypobaric hypoxia. We found that the immunoreactivity to the proteins studied in the hippocampus of rats was reduced in response to severe hypoxia. Hypoxic postconditioning sessions of mild hypobaric hypoxia (360 mmHg, 2 h, three times at intervals of 24 h) up regulated the expression of HIF-1α and erythropoietin in hippocampal CA1 neurons of rats that survived after severe hypoxia. Our results indicate that postconditioning led to compensation of hypoxia-induced neuron damage in the brain, which actively involved HIF-1α and erythropoietin.

Effects of Hypoxic Preconditioning on Expression of Transcription Factor NGFI-A in the Rat Brain after Unavoidable Stress in the “Learned Helplessness” Model

We report here our immunocytochemical studies establishing that the development of a depression-like state in rats following unavoidable stress in a “learned helplessness” model is accompanied by stable activation of the expression of transcription factor NGFI-A in the dorsal hippocampus (field CA1) and the magnocellular paraventricular nucleus of the hypothalamus, along with an early wave of post-stress expression, which died down rapidly, in the ventral hippocampus (the dentate gyrus) and a long period of up to five days of high-level expression in the neocortex. In rats subjected to three sessions of preconditioning consisting of moderate hypobaric hypoxia (360 mmHg, 2 h, with intervals of 24 h), which did not form depression in these circumstances, there were significant changes in the dynamics of immunoreactive protein content in the hippocampus, with a stable increase in expression in the ventral hippocampus and only transient and delayed (by five days) expression in field CA1. In the neocortex (layer II), preconditioning eliminated the effects of stress, preventing prolongation of the first wave of NGFI-A expression to five days, while in the magnocellular hypothalamus, conversely, preconditioning stimulated a second (delayed) wave of the expression of this transcription factor. The pattern of NGFI-A expression in the hippocampus, neocortex, and hypothalamus seen in non-preconditioned rats appears to reflect the pathological reaction to aversive stress, which, rather than adaptation, produced depressive disorders. Post-stress modification of the expression of the product of the early gene NGFI-A in the brain induced by hypoxic preconditioning probably plays an important role in increased tolerance to severe psychoemotional stresses and is an important component of antidepressant mechanisms.

The neurotrophin BDNF is involved in the development and prevention of stress-induced psychopathologies

The neurotrophin BDNF plays an important role in synaptic plasticity, the survival of neurons, and obviously, in the stress response of the body. We studied the time course of the changes in BDNF expression in the hippocampus and neocortex of rats after their exposure to pathogenic stress in models of endogenous depression and post-traumatic stress disorder (PTSD) and after treatment with hypoxic preconditioning, which prevented the development of anxiety–depressive conditions. Using immunohistochemistry we revealed that the development of experimental anxiety–depressive conditions was accompanied by a significant and stable decrease in the BDNF contents in hippocampal and neocortical neurons. In a model of depressive pathology, hypoxic preconditioning normalized the number of BDNF-expressing neurons early after stress exposure. In a model of PTSD, hypoxic preconditioning substantially and protractedly stimulated BDNF expression in the studied brain structures. Thus, the insufficient expression of this factor in the higher brain structures evidently reflected a maladaptive response to severe stress and may be a general feature of the pathogenesis of both depression and PTSD, whereas the significant and stable upregulation of BDNF expression is involved in the mechanisms of the anxiolytic effects of preconditioning and is important for the prevention of the development of PTSD.

The adaptive role of the CREB and NF-κB neuronal transcription factors in post-stress psychopathology models in rats

We studied the dynamics of the activation of the CREB and NF-κB transcription factors in areas of the rat brain after exposure to pathogenic psycho-emotional stress in the model of endogenous depression and the model of posttraumatic stress disorder (PTSD), as well as after application of hypoxic preconditioning, which prevents the formation of anxiety-depressive pathologies in these models. The development of anxiety-depressive pathology in both models was associated with reduced (10 times lower than the control maximum) or basal levels of the activating transcription factors CREB and NF-κB in the hippocampus and the neocortex. However, in the hypothalamus, NF-κB overactivation (up to an 18-fold increase above the control level) was detected in the model of depression. Therefore the lack of activation of these factors in the extra-hypothalamic areas of the brain may be a common component of the pathogenesis of both depression and PTSD, while NF-κB overactivation in the neurosecretory centers of the hypothalamus is obviously involved in the development of depressive conditions. Antidepressant and anxiolytic actions of hypoxic preconditioning were accompanied by a mild 2- to 6-fold increase in CREB and NF-κB levels in the neurons of the brain in both stress paradigms. Obviously, the maintenance of optimum activity of transcription factors in the neurons of the brain plays an important role in nonspecific compensatory processes that enhance the adaptive potential of the brain under stress conditions.

Involvement of the transcription factor c-Fos in the protective effect of hypoxic preconditioning in a model of post-traumatic stress disorder in rats

A quantitative immunocytochemical method was used to study the dynamics of the expression of the transcription factor c-Fos in the hypothalamus, hippocampus, and neocortex during the development of anxiety in the post-traumatic stress disorder (PTSD) model in control rats and rats preconditioned by hypoxia, which is known to have an anxiolytic effect. It was found that the induction of experimental PTSD was associated with a significant and stable increase in the level of c-Fos in all investigated brain areas. The maximum increase according to amplitude (up to 30 times) and duration (about 10 days) was observed in the hypothalamus. This overexpression may be associated with corticotropin-releasing hormone hyperproduction, which is typical of PTSD. In animals that were subjected to triple hypoxic preconditioning with mild repetitive hypobaric hypoxia (360 mm Hg for 2 hours daily, for 3 days), which prevented the traumatic stress-induced pathology, c-Fos overexpression was completely or partially blocked. These data indicate that stable overexpression of the c-Fos transcription factor in the neocortex, hippocampus, and hypothalamus apparently participates in the mechanisms of PTSD development, whereas its blockade plays an important role in increasing the brain’s tolerance to stresses and in protection against stress-induced pathologies.

Characteristics of the transcription factor HIF-1α expression in the rat brain during the development of a depressive state and the antidepressive effects of hypoxic preconditioning

The dynamics of HIF-1α expression during the development of stress-related depression, as well as after hypoxic preconditioning (HP), which has an antidepressant-like effect, were studied in the hippocampus, paraventricular hypothalamic nucleus, and neocortex of rats, using an immunocytochemical method. It has been found that the factor HIF-1α is induced in neurons in response to psychoemotional stress that causes the development of experimental depression in rats in the “learned helplessness” model. The profile of the stress-induced expression of HIF-1α in the hippocampus has a two-wave character: early expression on the first day and the delayed expression 10 days after the stress. No significant change was found in the neocortex. In the hypothalamus, up-regulation of HIF-1α expression was delayed (5–10 days). After HP by a moderate repetitive hypobaric hypoxia, which prevents the development of the depressive state in rats, the post-stress expression of HIF-1α was considerably altered in the brain regions studied. In the hippocampus of HP rats, the peak of the early expression lasted for about 5 days after the stress; we observed a multifold increase in its amplitude. In contrast, the HIF-1α delayed peak was eliminated. A similar but smaller effect of HP was also observed in the hypothalamus. The data obtained indicate that delayed HIF-1α expression in the hippocampus and hypothalamus was apparently involved in the mechanisms of pathogenesis of the depressive pathology. However, strong modifications in early and late post-stress expression of HIF-1α caused by HP obviously play an important role in increasing the brain’s tolerance to severe stresses and protection against the development of stress-induced depressive pathologies.

The Anxiolytic Effects of Moderate Hypoxia and Remote Ischemia in the Posttraumatic Stress Disorder Model Are Accompanied by Modification of Functioning of the Hypothalamic–Pituitary–Adrenal Axis

Using a model of posttraumatic stress disorder (PTSD) in rats, it has been shown that conditioning by moderate hypobaric hypoxia (360 mmHg, three times for 2 h with 24-h interval) or limb ischemia–reperfusion (three times for 5 min with 15-min interval) prevents premature suppression of release of the stress hormone corticosterone to the blood plasma, which is typical of the triggering of pathological fast negative feedback of the hypothalamic–pituitary–adrenal axis (HPA) in this pathology. Hypoxic or remote ischemic preand postconditioning also increased the basal level of this hormone, which is significantly reduced during the formation of experimental PTSD. Thus, the pronounced anxiolytic effect of these conditioning types in the PTSD model may be mediated by the normalization of HPA regulation by a feedback mechanism and prevention of a decrease in its basal activity. Further research on the decoding of neurohumoral mechanisms of the stress-protective action of conditioning will speed up the introduction of these effective non-drug ways of prevention and correction of stress-induced pathologies into medical practice.

The dynamics of HIF-1α expression in the rat brain at different stages of experimental posttraumatic stress disorder and its correction with moderate hypoxia

The dynamics of the expression of the HIF factor-1 α-subunit, which is related to products of early genes, has been studied in the neocortex, the hippocampus, and the hypothalamus of rats during the development of posttraumatic stress disorder (PTSD) in a stress–restress model and using triple moderate hypobaric hypoxia (MH3), which prevents the formation of this anxiety pathology. The immunohistochemical method has shown that after pathogenic traumatic stress (TS), during the primary (“hidden”) period of modeled PTSD formation, the level of HIF-1α expression did not change significantly; however, after restress it rapidly increased in all regions of the brain. An increased expression of this factor remained in animals with experimental PTSD for up to 10 days after restress. Exposure to MH3 before TS or before restress compensated for these disturbances: fully in the hippocampus and partly in the neocortex; it inhibited the prolonged over-induction of the HIF-1α, which may be one of the mechanisms that mediate an anxiolytic effect of hypoxia. Along with this, preconditioning with MH3 significantly decreased the content of HIF-1α after TS, thus preventing activation of the HIF-1 factor during the hidden period, which is likely associated with the formation of pathological reactivity to restress. These facts indicate the pathogenetic role of HIF-1 in certain periods of experimental PTSD and the correcting effect of hypoxic preconditioning.