Modulating Effect of Cytokines on Mechanisms of Synaptic Plasticity in the Brain.

After accumulation of data showing that resident brain cells (neurons, astrocytes, and microglia) produce mediators of the immune system, such as cytokines and their receptors under normal physiological conditions, a critical need emerged for investigating the role of these mediators in cognitive processes. The major problem for understanding the functional role of cytokines in the mechanisms of synaptic plasticity, de novo neurogenesis, and learning and memory is the small number of investigated cytokines. Existing concepts are based on data from just three proinflammatory cytokines: interleukin-1 beta, interleukin-6, and tumor necrosis factor-alpha. The amount of information in the literature on the functional role of antiinflammatory cytokines in the mechanisms of synaptic plasticity and cognitive functions of mature mammalian brain is dismally low. However, they are of principle importance for understanding the mechanisms of local information processing in the brain, since they modulate the activity of individual cells and local neural networks, being able to reconstruct the processes of synaptic plasticity and intercellular communication, in general, depending on the local ratio of the levels of different cytokines in certain areas of the brain. Understanding the functional role of cytokines in cellular mechanisms of information processing and storage in the brain would allow developing preventive and therapeutic means for the treatment of neuropathologies related to impairment of these mechanisms.

Effect of Audiogenic Seizures on the Dynamics of Fatty Acid Composition of Hippocampal Dental Gyrus in Krushinsky-Molodkina Rats.

Changes in the fatty acid composition of the hippocampal dentate gyrus in Krushinsky-Molodkina rats with hereditary predisposition to audiogenic seizures were studied in 1, 3, and 14 days after 1 or 5 seizures. Seizure activity changed the content of saturated and monounsaturated as well as polyunsaturated fatty acids at different terms after seizures. After seizures, the content of individual fatty acids changed in different directions. Similar shifts after 1 and 5 seizures were observed only for eicosapentaenoic acid at all observation terms. More pronounced changes in fatty acid composition were observed after 5 seizures. These results can be useful for the development of new approaches to correction of seizure activity.

Role of BK(Ca) Potassium Channels in the Mechanisms of Modulatory Effects of IL-10 on Hypoxia-Induced Changes in Activity of Hippocampal Neurons.

We studied the contribution of large conductance Ca(2+)-activated potassium channels (BKCa) in the mechanisms of neuromodulatory effects of anti-inflammatory cytokine IL-10 on hypoxiainduced changes in activity of CA1 pyramidal neurons in rat hippocampus. We used the method of registration of population spikes from CA1 pyramidal neurons in hippocampal slices before, during, and after exposure to short-term episodes of hypoxia. Selective blocker (iberiotoxin) and selective activator of BKCa (BMS-191011) were used to evaluate the contribution of these channels in the mechanisms of suppressive effects of IL-10 on changes in neuronal activity during hypoxia and development of post-hypoxic hyperexcitability. It was shown that BKCa are involved in the modulatory effects of IL-10 on hypoxia-induced suppression of activity of CA1 pyramidal neurons in the hippocampus and development of post-hypoxic hyperexcitability in these neurons.

[THE CHANGES IN FATTY ACIDS COMPOSITION IN THE AUDITORY CORTEX OF RATS AFTER A SINGLE AUDIOGENIC SEIZURE].

We found small but statistically significant increase in the number of stable to peroxide oxidation saturated and monounsaturated fatty acids in the auditory cortex of KM rats in comparison with control Wistar ones. The levels of fatty acids in the cells of the auditory cortex of KM rats were studied at different times (1 h, 1 day, 3 days and 14 days) after a single audiogenic seizure. The changes in fatty acids composition in auditory cortex of KM rats were found already in time point 1 h after convulsion, the maximal decrease of fatty acids levels was observed at 3 days after convulsion. These data suggest that the fatty acids pool in this time was depleted. Finally, we found the recovery of the better part of fatty acids in the auditory cortex of KM rats to 14 day after convulsion. These results can be used for development of new approaches to eliminate brain damage after seizures.

Memory disorders in rats after impairment of the dorsal hippocampal CA3 field with kainic acid.

We studied the possibility of conditioning of food-procuring response in animals after impairment of the dorsal hippocampal region with kainic acid. Histological studies of brain sections showed that the greater part of dorsal hippocampal CA3 pyramidal neurons were lyzed in 2 weeks after kainic acid microinjection into the hippocampus. Morphological signs of necrosis were found in CA2 and CA4 neurons at the interface with CA3 field. Preinjection of anti-inflammatory cytokine IL-10 virtually did not prevent neuronal death. Damage to the hippocampus impaired learning of experimental animals and they required more attempts for reliable performance of the skill than the controls. Short-term memory of experimental rats was retained, which was proven by the results of single-day training. It was hypothesized that memory dysfunctions in animals after damage to the dorsal hippocampal CA3 field and to the hippocampus in general are determined by impaired transition of short-term into long-term memory.

Comparison of effects of ATP-gated potassium channel blockers on activity variations of rat CA1 pyramidal neurons in hippocampal slices triggered by short-term hypoxia.

The study compared the effects of KATP channels blockers 5-hydroxydecanoat and glibenclamide on rapid hypoxic preconditioning and posthypoxic hyperexcitability of CA1 pyramidal neurons in rat hippocampal slices induced by short-term hypoxia. The population spikes of CA1 neurons were recorded before, during, and after exposure to a short-term hypoxia. The blockers of KATP channels significantly degraded the potency of hypoxia episodes to inhibit the evoked neuronal population activity. In contrast to glibenclamide, 5-hydroxydecanoat eliminated the preconditioning action of hypoxia. Despite mitochondrial KATP channels play an important role in the mechanisms of rapid hypoxic preconditioning in hippocampal CA1 pyramidal neurons, the tested KATP channels blockers produced no significant effect on the development of posthypoxic hyperexcitability in these neurons.

[Effect of seizure activity on subunit composition of Ca2+/calmodulin-dependent protein kinase II in hippocampus of Krushinskii-Molodkina rats].

Using a Western blot method it was been shown that a aontent of beta-CaMKII subunit was decreased on 40 % in hippocampus of Krushinskii-Molodkina rats (rats, genetically prone to audiogenic seizures) in comparison with normal Wistar rats. Additionally, we have investigate the temporal modifications of alpha/beta-CaMKII subunits in dependent from number audiogenic fits (n = 1 or n = 5). The level of beta-CaMKII in hippocampus of naive KM rats and KM rats in 24 h after single audiogenic seizure was not different, but it was increased on 56 % through 72 h after convulsion fit. In contrast, both 5-fold seizures and 20-fold seizures (audiogenic kindling) reduced the beta-CaMKII levels in comparison with naive group. The levels of alpha-subunit protein kinase in hippocampus these experimental groups of animals were not different. We suggest a hypothesis, that modifications alpha/beta ratio can be viewed as specific "homeostatic mechanism", which provide for neuronal function in hippocampus, thereby changing the rules governing synaptic plasticity in dependence from preliminary neuronal activity.

[Fullerenes C60, antiamyloid action, the brain and cognitive processes].

A short review of investigations along a new line: the antiamyloid action of fullerenes C60 and correction of disturbed cognitive processes is presented. The prospects for the development of drugs based on fullerenes acting on the key molecular mechanisms at the early stage of Alzheimer's disease are discussed.

[Role of Ca(2+)-activated potassium channels with high-conductivity in hypoxic preconditioning and posthypoxic hyperexcitability in CA1 hippocampal neurons in vitro].

The role of highly conductive Ca(2+)-activated potassium channels (BK(Ca)-channels) of rat hippocampal CA1 neurons in the processes induced by repeated episodes of brief hypoxia was investigated. A specific blocker of BK(Ca)-channels iberiotoxin (10-20 nM) tended to decrease the efficiency of hypoxia in depressing the amplitude of population spikes during hypoxic episode. Iberiotoxin significantly abolished both the preconditioning effect of hypoxic episodes and the posthypoxic hyperexcitability of pyramidal neurons. It is suggested that BK(Ca)-channels play an important role in the mechanisms of such forms of neuroplasticity as rapid hypoxic preconditioning and posthypoxic hyperexcitability in hippocampal CA1 neurons.

[Single audiogenic seizure promotes the increased levels of calcineurin and Ca(2+)-calmodulin-dependent protein kinase II in the rat brain].

Ca(2+)-calmodulin-dependent protein kinase II (CaMKII) and protein phosphatase 2B (calcineurin) play a critical role in modulation responses of nerve cells to Ca(2+)-signal. Here we asked the question, whether and how these enzymes may become affected by single seizure activity. Male epilepsy-prone Krushinsky-Molodkina rats were exposed to single sound stimulation (80 dB, 12-15 kHz). Biochemical studies carried out two days after the sound exposure. Immunoblots of hippocampal and cortical (from sensomotor area) homogenates reacted with monoclonal antibodies to neurospecific alpha-subunit CaMKII showed an increased presence of this protein in seizured animals in comparison with naive controls. The level of the calcineurin catalytic subunit was increased in the hippocampus only. Additionally, studies of CaMKII activity revealed that the total enzyme activity from hippocampus and cortex of seizured rats was increased as compared with controls. However, it was no differences in functional (Ca(2+)-calmodulin-independent) CaMKII activity between experimental and control groups. It was suggested that observed long-lasting changes in rats brain induced by seizure activity may be a one in a number adaptative mechanisms against neuronal exitability.

The effects of interleukin-10 on the development of epileptiform activity in the hippocampus induced by transient hypoxia, bicuculline, and electrical kindling.

The comparative effects of the anti-inflammatory cytokine interleukin-10 on the development of epileptiform activity were studied in hippocampal field CA1 neurons in different models of epileptogenesis not accompanied by visible morphological lesions in brain cells: 1) a model of hypoxic kindling in rat hippocampal slices; 2) a disinhibitory model of epileptogenesis in rat hippocampal slices using the GABAA receptor blocker bicuculline; and 3) a partial electrical kindling model in intact rats. Interleukin-10 (1 ng/ml) blocked the development of post-hypoxic hyperexcitability of field CA1 pyramidal neurons in hippocampal slices, decreasing the effectiveness of hypoxia in suppressing neuron activity during the hypoxic episode. Interleukin-10 had no effect on the initiation of epileptiform activity in pyramidal neurons induced by the proconvulsant bicuculline. Single intrahippocampal injections of interleukin-10 at a dose of 1 ng in 5 microl suppressed the development of focal convulsions ("ictal" discharges) at the stimulation site in partial kindling in freely moving animals for several hours after administration. However, this cytokine had no effect on the duration of the "interictal" component of focal afterdischarges or on the severity of behavioral seizures. These results show that the anti-inflammatory cytokine interleukin-10, at the concentrations used here, has not only antihypoxic activity, but also a protective effect in relation to the initiation of the "ictal," but not the "interictal" component of epileptiform activity in hippocampal neurons.

Partial kindling induces neurogenesis, activates astrocytes and alters synaptic morphology in the dentate gyrus of freely moving adult rats.

A partial kindling procedure was used to investigate the correlation between focal seizure development and changes in dendritic spine morphology, ongoing neurogenesis and reactive astrogliosis in the adult rat dentate gyrus (DG). The processes of neurogenesis and astrogliosis were investigated using markers for doublecortin (DCX), 5-bromo-2-deoxyuridine (BrdU) and glial fibrillary acidic protein (GFAP). Our data demonstrate that mild focal seizures induce a complex series of cellular events in the DG one day after cessation of partial rapid kindling stimulation consisting (in comparison to control animals that were electrode implanted but unkindled), firstly, of an increase in the number of postmitotic BrdU labeled cells, and secondly, an increase in the number of DCX labeled cells, mainly in subgranular zone. Ultrastructural changes were examined using qualitative electron microscope analysis and 3-D reconstructions of both dendritic spines and postsynaptic densities. Typical features of kindling in comparison to control tissue included translocation of mitochondria to the base of the dendritic spine stalks; a migration of multivesicular bodies into mushroom dendritic spines, and most notably formation of "giant" spinules originating from the head of the spines of DG neurons. These morphological alterations arise at seizure stages 2-3 (focal seizures) in the absence of signs of the severe generalized seizures that are generally recognized as potentially harmful for neuronal cells. We suggest that an increase in ongoing neurogenesis, reactive astrogliosis and dendritic spine reorganization in the DG is the crucial step in the chain of events leading to the progressive development of seizure susceptibility in hippocampal circuits.

[Effects of interleukin-10 on the epileptiform activity development in the hippocampus induced by brief hypoxia, bicuculline and electrical kindling].

The comparative effects of antiinflammatory cytokine interleukin-10 on the epileptiform activity development in CA1 hippocampal neurons were studied in different functional models of epileptogenesis that are not accompanied the visible morphological disturbances in the brain cells: --in vitro hypoxic model in the rat hippocampal slices; 2--in vitro disinhibitory model with using GABAA antagonist, bicuculline, in the rat hippocampal slices; 3--partial hippocampal kindling model in freely moving rats. Interleukin-10 (1 ng/ml) depressed the posthypoxic hyperexcitability in CA1 pyramidal neurons of the rat hippocampal slices through a decrease of the effectiveness of hypoxia to depresses the functional neuronal activity in the rat hippocampal slices during hypoxic episode. On the other hand, interleukin-10 (1 ng/ml) did not affect an initiation of epileptiform activity in CA1 pyramidal neurons of the rat hippocampal slices induced by bicuculline. Interleukin-10 (1 ng/5 microl) applied to the dorsal hippocampus in awake rats depressed an initiation of focal seizures ("ictal"-like components of afterdischarges) induced by hippocampal kindling during the first six hours after an application. However, this cytokine did not affect neither the duration of "interictal"-like component of afterdischarges nor motor seizure development. Thus, our findings showed that antiinflammatory cytokine interleukin-10, in addition to its antihypoxic action, exert the neuroprotective effect on the initiation of "ictal"-like, but not "interictal"-like, epileptiform discharges.

[The role of cytokines in the seizure activity development in the brain].

In this review, the role of pro-inflammatory cytokines, interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-alpha), and anti-inflammatory cytokine, IL-10, in the seizure activity development is analyzed. In recent years, there has been increasing evidence that the transformation of normal pattern of neuronal activity to paroxysmal one is associated with the increased production of these cytokines in the brain. However, the present results indicate that expressions of IL-1, TNF-alpha and IL-10 in the brain are associated with cell injury rather than with seizures per se. These findings suggest that, in response to seizures, these cytokines cause both neuroprotective or neurodegenerative effects and, as a consequence of these effects, the further facilitation or depression of seizure activity.

Protective effects of interleukin-10 on the development of epileptiform activity evoked by transient episodes of hypoxia in rat hippocampal slices.

The aim of the present work was to study the effects of interleukin-10 at concentrations of 1 and 10 ng/ml on the development of epileptiform discharges evoked in pyramidal neurons in field CA3 in rat hippocampal slices by transient episodes of hypoxia. Three 3-min episodes of hypoxia led to decreases in the generation threshold for evoked trains of population spikes and an increase in the number of population spikes per train in pyramidal neurons of field CA1. Interleukin-10 at a concentration of 1 ng/ml completely eliminated the development of epileptiform activity, while its protective effect was less marked at a concentration of 10 ng/ml. These effects of interleukin-10 on living hippocampal slices in in vitro conditions show that they may be associated with the functions of this cytokine as an intercellular mediator of the central nervous system itself rather than being mediated by the peripheral immune system. The results of these studies provide the first experimental evidence of the action of the anti-inflammatory cytokine interleukin-10 on the development of hypoxia-evoked epileptiform events in the hippocampus.

[Protective effect of interleukin-10 on the development of epileptiform activity induced by brief hypoxic episodes in the rat hippocampal slices].

The aim of the study was to investigate the effect of interleukin-10 (IL-10) (1 and 10 ng/ml) on the development of epileptiform activity induced by brief hypoxic episodes in CA1 area of rat hippocampal slices. Three three-minute hypoxic episodes induced a sustained decrease in the threshold of evoked population spike (PS) burst and an increase in the number of PSs in the PS response. IL-10 (1 ng/ml) completely abolished the development of epileptiform activity whereas the effect of IL-10 (10 ng/ml) was weaker. The protective effect of IL-10 on the hyperexcitability of the local neuronal network in hippocampal slices indicate that this cytokine can function as an intercellular mediator in the brain. The present results are the first experimental evidence of a protective role of anti-inflammatory IL-10 in the development of epileptiform events induced by brief episodes of hypoxia in the hippocampus.