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The composition of biologically active substances in the aqueous total fraction of the 95% EtOH extract of the aerial part of Alfredia cernua (L.) Cass. possessing antiamnesic properties was studied using chromatographic and mass-spectrometric methods. The total amount of N-containing compounds in the fraction was found to be 10.63 ± 1.65%. Fifteen amino acids (1.99 ± 0.11%), seven of which (0.76 ± 0.012%) were essential and included lysine, threonine, valine, isoleucine, leucine, phenylalanine, and histidine, were identified using an amino-acid analyzer. The total fraction was dominated by glutamic and aspartic acids, valine, proline, alanine, and histidine. The composition included a total of 43 elements (1.35 ± 0.1%), of which 19 were essential or conditionally essential. The predominant elements were K, Mg, Zn, Rb, P, Ca, and Mn. Carbohydrates dominated (32 ± 3.1%) the fraction.

Barriergenesis is the process of maturation of the primary vascular network of the brain responsible for the establishment of the blood-brain barrier. It represents a combination of factors that, on the one hand, contribute to the process of migration and tubulogenesis of endothelial cells (angiogenesis), on the other hand, contribute to the formation of new connections between endothelial cells and other elements of the neurovascular unit. Astrocytes play a key role in barriergenesis, however, mechanisms of their action are still poorly examined. We have studied the effects of HIF-1 modulators acting on the cells of non-endothelial origin (neurons and astrocytes) on the development of the blood-brain barrier in vitro. Application of FM19G11 regulating expression of HIF-1 activity and GSI-1 suppressing gamma-secretase and/or proteasomal activity resulted in the elevated expression of thrombospondins and matrix metalloproteinases in the developing blood-brain barrier. However, it caused the opposite effect on VEGF expression thus promoting barrier maturation in vitro.

Все анатомические элементы, которые входят в гематоэнцефалический барьер (ГЭБ), играют важную роль в регуляции проницаемости и гомеостаза ЦНС в норме и патологии. Этими элементами являются эндотелиальные клетки, перициты, астроглия и нейроны, и все они входят в понятие «нейроваскуляторная единица» (НВЕ). Будучи интеграционной системой, НВЕ тонко регулирует синаптическую пластичность нейронов, нейрогенез, межклеточные взаимодействия и проницаемость ГЭБ. Эндотелиальные клетки капилляров головного мозга являются важной составляющей НВЕ. В этом обзоре мы обсуждаем значительную роль эндотелиальных клеток капилляров мозга в поддержании структурной и функциональной целостности ГЭБ. В последние десятилетия большое внимание было уделено анализу экспрессии белков тесных контактов и белков адгезионных контактов в эндотелиальных клетках капилляров мозга и лишь относительно небольшое число исследований было сфокусировано на оценке экспрессии и функциональной активности ионных каналов в этих клетках, несмотря на то, что существует все большее число доказательств их важной роли в регуляции функций НВЕ/ГЭБ. В целом электрофизиологические свойства эндотелиальных клеток капилляров зависят от экспрессии различных ионных каналов, чья активность, по всей вероятности, координирует некоторые виды межклеточных взаимодействий в НВЕ и клетках артериол головного мозга. Мы остановили свое внимание на роли ионных каналов в регуляции активности клеток НВЕ, гладкомышечных клеток артериол и в модуляции локального кровотока головного мозга. Большое место в обзоре отведено лиганд-регулируемым ионным каналам, каналам, регулируемым внутриклеточными кальциевыми депо, TRP-каналам, кальций-активируемым и потенциалзависимым калиевым каналам в эндотелиальных клетках капилляров мозга и клетках артериол головного мозга. Понимание роли ионных каналов в контроле церебрального кровотока позволит определить новые терапевтические мишени для восстановления функциональной целостности НВЕ/ГЭБ при различных патологических состояниях (ишемия, нейровоспаление, нейродегенерация) in vivo и в моделях ГЭБ in vitro.All anatomical elements contributing to the blood-brain barrier (BBB) play a crucial role in maintaining the permeability and CNS homeostasis under physiological/pathological conditions. These elements are endothelial cells, pericytes, astroglia, and neurons that are known as a neurovascular unit (NVU). Being the integral system, NVU contributes to the regulation of neuroplasticity, neurogenesis, intercellular communications and permeability of BBB. Brain capillary endothelial cells (BCEC) are the very important part of NVU. In this review, we discuss the critical role of BCEC ion channels in BBB structural and functional integrity. In last decades, much attention has been paid to the expression of tight junctions and adherence junctions in BCEC whereas less number of studies was focused on the expression and functioning of ion channels in BCEC, however, there is growing evidence supporting their important role in the regulation of NVU/BBB functions. In general, electrophysiological properties of BCEC depend on the expression of various ion channels whose activity, presumably, coordinates intercellular communication within the NVU. Particularly, we focus on BCEC ion channels-dependent mechanisms of NVU functioning, arteriole smooth muscle cells dynamic modulation, and changes in the regional cerebral blood flow. We put special attention on ligand-gated ion channels, store-operated calcium channels, TRP ion channels, calcium-activated, voltage-gated potassium channels in BCEC. Understanding the role of ion channel signaling in the control of cerebral blood flow will helps to reveal the potential therapeutic targets to recover the NVU/BBB functional integrity in different pathological conditions (ischemia, neuroinflammation, neurodegeneration) both in vivo and in vitro BBB models.

An important mechanism of neuronal plasticity is neurogenesis, which occurs during the embryonic period, forming the brain and its structure, and in the postnatal period, providing repair processes and participating in the mechanisms of memory consolidation. Adult neurogenesis in mammals, including humans, is limited in two specific brain areas, the lateral walls of the lateral ventricles (subventricular zone) and the granular layer of the dentate gyrus of the hippocampus (subgranular zone). Neural stem cells (NSC), self-renewing, multipotent progenitor cells, are formed in these zones. Neural stem cells are capable of differentiating into the basic cell types of the nervous system. In addition, NSC may have neurogenic features and non-specific non-neurogenic functions aimed at maintaining the homeostasis of the brain. The microenvironment formed in neurogenic niches has importance maintaining populations of NSC and regulating differentiation into neural or glial cells via cell-to-cell interactions and microenvironmental signals. The vascular microenvironment in neurogenic niches are integrated by signaling molecules secreted from endothelial cells in the blood vessels of the brain or by direct contact with these cells. Accumulation of astrocytes in neurogenic niches if also of importance and leads to activation of neurogenesis. Dysregulation of neurogenesis contributes to the formation of neurological deficits observed in neurodegenerative diseases. Targeting regulation of neurogenesis could be the basis of new protocols of neuroregeneration.

The formation and functional plasticity of the blood–brain barrier are inextricably linked to the molecular events occurring in cerebral neurovascular unit during the embryonic and early postnatal development of the organism. To study the features of the barrier genesis under physiological conditions, as well as after perinatal hypoxia and stress in early life, tight junction proteins of cerebral endothelial cells (the number of JAM-, ZO1-, and CLDN5-positive cells) in rats at the age of 7 (P7), 28 (P28), and 70 (P70) days were investigated. It was found that, under physiological conditions, the number of cells expressing JAM, ZO1, and CLDN5 slightly increase in the period from P7 to P70 in the cortex, hippocampus, and amygdala of the brain. After perinatal hypoxia, the number of cells expressing the proteins of tight junctions (JAM, CLDN5) is significantly increased to the age of P28–P70, while the number of ZO1+ cells in the same period of time is reduced. Early life stress causes an imbalance between the expression of ZO1 and other proteins of tight junctions, but these changes are opposite in direction.

Patch clamp is a golden standard for studying (patho)physiological processes affecting membranes of excitable cells. This method is rather labor-intensive and requires well-trained professionals and long-lasting experimental procedures; therefore, accurate designing of the experiments with patch clamp methodology as well as collecting and analyzing the data obtained are essential for the widely spread implementation of this method into the routine research practice. Recently, the method became very prospective not only for the characterization of single excitable cells but also for the detailed assessment of intercellular communication, i.e. within the neurovascular unit. Here, we analyze the main advantages and disadvantages of patch clamp method, with special focus on the tendencies in clamping technique improvement with the help of patch electrodes for the assessment of intercellular communication in the brain.

Formation and functional plasticity of the blood-brain barrier is associated with the molecular events that occur in the brain neurovascular unit in the embryonic and early postnatal development. To study the characteristics of barriergenesis under physiological conditions, as well as recovering from perinatal hypoxia and early life stress, we examined the expression of proteins of cerebral endothelial tight junctions (JAM, ZO1, CLDN5) in rats aged 7, 28 and 70 days of postnatal development (P7—P70). Under physiological conditions, we have found that the number of endothelial cells expressing JAM, ZO1, CLDN5 slightly increases in the cortex, hippocampus and amygdala of the brain in the period from P7 to P70. Perinatal hypoxia significantly increased the number of cells expressing proteins of tight junction proteins (JAM, CLDN5) up to the age P28—P70, whereas the number of cells expressing ZO1 was reduced in the same period of time. Early life stress led to an imbalance between the number of cells expressing ZO1 proteins and that expressing tight junctions proteins, but these changes were in opposite direction to that observed in perinatal hypoxia

Metabolic activity of cells within a neurovascular unit is among the factors determining structural and functional integritY of the blood-brain barrier and the an- giogenesis process. in order to verify the hypothesis about the role Of g1YcolYtic activity in the perivascula astroglialcells associated with lactate release in the development of functioning of cerebral microvessel endothelial cells, we have used a three-component model of the brain neurovascular unit in vitro. The cells o f n o n -en d o th elia l o rig in w ere in c u b a te d in th e p rese n ce o f m o d u la to rs o f la c ta te pro d u c n ago ni glu c ose ta a G ly c o s o) , bas t h e oe t a n t a at- blocker of monocarboxylate transporters MCTlprCT and recepltiors of3Ctate0produasan (2-donisyoflactate G e8 breceptor) Iasa estbishe vthat that te suppression of lactate production and transport, prdc o1,adrcpin(C-O-Aa n (2gdoxysgflucoase as a glycolysis inhibitor), transport (phloretin as a sukr of lacaroduto transport , aswellasastimultionof3lactate receptors in astroglial cells, lead to aberrant development of endothelial layer, ther by u g g e tin t h efor atio o f anti ngi gencmi roen ircm ent for cerebral endothelium due to inappropriate lactate-m ediated effects. KeYw.ords:-n-eur-ovascular unit; metabolism; glYcolysis; lactate.

Barriergenesis is the process of maturation of the primary vascular network of the brain responsible for the establishment of the blood-brain barrier. It represents a combination of factors that, on the one hand, contribute to the process of migration and tubulogenesis of endothelial cells (angiogenesis), on the other hand, contribute to the formation of new connections between endothelial cells and other elements of the neurovascular unit. Astrocytes play a key role in barriergenesis, however, mechanisms of their action are still poorly examined. We have studied the effects of HIF-1 modulators acting on the cells of non-endothelial origin (neurons and astrocytes) on the development of the blood-brain barrier in vitro. Application of FM19G11 regulating expression of HIF-1 activity and GSI-1 suppressing gamma-secretase and/or proteasomal activity resulted in the elevated expression of thrombospondins and matrix metalloproteinases in the developing blood-brain barrier. However, it caused the opposite effect on VEGF expression thus promoting barrier maturation in vitro.Bar'erogenez – éto protsess sozrevaniia pervichnoĭ sosudistoĭ seti golovnogo mozga, pri kotorom proiskhodit stanovlenie gematoéntsefalicheskogo bar'era. On predstavliaet soboĭ sochetanie faktorov, kotorye, s odnoĭ storony, sposobstvuiut protsessu migratsii i tubulogenezu éndotelial'nykh kletok (angiogenez), s drugoĭ storony, sposobstvuiut formirovaniiu novykh sviazeĭ mezhdu éndoteliotsitami i drugimi élementami neĭrovaskuliarnoĭ edinitsy. Perivaskuliarnye astrotsity vnosiat znachitel'nyĭ vklad v bar'erogenez, odnako mekhanizmy étogo uchastiia malo izucheny. V rabote issledovano deĭstvie moduliatorov HIF-1 v kletkakh neéndotelial'noĭ prirody (neĭrony i astrogliia) na formirovanie gematoéntsefalicheskogo bar'era in vitro. Primenenie FM19G11, reguliruiushchego ékspressiiu HIF-1, i GSI-I, podavliaiushchego aktivnost' gamma-sekretazy i/ili proteasom, privodit k uvelicheniiu ékspressii trombospondinov i matriksnykh metalloproteinaz v razvivaiushchemsia gematoéntsefalicheskom bar'ere, no okazyvaet raznonapravlennyĭ éffekt v otnoshenii produktsii VEGF, chto, v tselom, sposobstvuet sozrevaniiu bar'era in vitro.

Neurodegeneration is progressive neuronal death. Research on neuroinflammation mechanisms is necessary for understanding of the pathogenesis of acute and chronical neurodegeneration. The purpose of this research was to evaluate the coexpression of neuroinflammation molecules (CD157, CD11b/CD18) in the midbrain of animals under physiological conditions and in an experimental model of Parkinson’s disease. Immunohistochemical evaluation of the coexpression of CD157 and MAC-1 was performed in rat midbrain sections. A significant increase in the number of cells that coexpress CD157/MAC-1 (CD11b/CD18) was found in the midbrain during Parkinson’s disease. The possible roles of the neuroinflammatory molecules CD157 and MAC-1 in the pathogenesis of Parkinson’s disease may include activation of microglial cells and performance of the neuroinflammatory response.

We studied in vitro development of brain progenitor cells isolated from healthy 7-9-month-old Wistar rats and rats with experimental Alzheimer's disease kept under standard conditions and in enriched (multistimulus) environment in vivo. Progenitor cells from healthy animals more rapidly formed neurospheres. Considerable changes at the early stages of in vitro development of brain progenitor cells were observed in both groups kept in enriched environment.

The blood-brain barrier (BBB) consists of differentiated cells integrating in one ensemble to control transport processes between the central nervous system (CNS) and peripheral blood. Molecular organization of BBB affects the extracellular content and cell metabolism in the CNS. Developmental aspects of BBB attract much attention in recent years, and barriergenesis is currently recognized as a very important and complex mechanism of CNS development and maturation. Metabolic control of angiogenesis/barriergenesis may be provided by glucose utilization within the neurovascular unit (NVU). The role of glycolysis in the brain has been reconsidered recently, and it is recognized now not only as a process active in hypoxic conditions, but also as a mechanism affecting signal transduction, synaptic activity, and brain development. There is growing evidence that glycolysis-derived metabolites, particularly, lactate, affect barriergenesis and functioning of BBB. In the brain, lactate produced in astrocytes or endothelial cells can be transported to the extracellular space via monocarboxylate transporters (MCTs), and may act on the adjoining cells via specific lactate receptors. Astrocytes are one of the major sources of lactate production in the brain and significantly contribute to the regulation of BBB development and functioning. Active glycolysis in astrocytes is required for effective support of neuronal activity and angiogenesis, while endothelial cells regulate bioavailability of lactate for brain cells adjusting its bidirectional transport through the BBB. In this article, we review the current knowledge with regard to energy production in endothelial and astroglial cells within the NVU. In addition, we describe lactate-driven mechanisms and action of alternative products of glucose metabolism affecting BBB structural and functional integrity in developing and mature brain.

ИЗУЧЕНИЕ МЕТАБОЛИЧЕСКОГО СОПРЯЖЕНИЯ И МЕЖКЛЕТОЧНЫХ ВЗАИМОДЕЙСТВИЙ НА МОДЕЛИ НЕЙРОВАСКУЛЯРНОЙ ЕДИНИЦЫ IN VITRO А. В. Моргун, Н. В. Кувачева, Е. Д. Хилажева, Е. А. Пожиленкова, А. Б. Салмина ГБОУ ВПО Красноярский государственный медицинский университет имени проф. В. Ф. Войно-Ясенецкого Министерства здравоохранения РФ, ректор – д. м. н., проф. И. П. Артюхов; кафедра педиатрии ИПО, зав. – д. м. н., проф. Т. Е. Таранушенко; кафедра биохимии с курсами медицинской, фармацевтической и токсикологической химии, зав. – д. м. н., проф. А. Б. Салмина; НИИ молекулярной медицины и патобиохимии, руководитель – д. м. н., проф. А. Б. Салмина.

The review covers current concepts on cell and molecular mechanisms of neuroinflammation and aging with the special focus on the regulation of cytokine-producing activity of astroglial cells and intercellular communication. The review reflects that a key component of the aging phenomenon as a result of ineffective implementation of anti-inflammatory response are processes of the dysregulated cytokine production, in particular, an increase in the secretion of proinflammatory cytokines and an imbalance in the expression of the receptors and receptor associated proteins. Interpretation of the molecular mechanisms of cell conjugating neuroinflammation and aging cells can give rise to new therapeutic strategies that are relevant to the treatment of a wide range of central nervous system diseases and the development of new experimental models of diseases of the central nervous system.

The aim of this research was to assess a role of connexin 43 (Cx43) and associated molecule CD38 in the regulation of cell-cell interactions in the neurovascular unit (NVU) in vitro in physiological conditions and in hypoxia.Materials and methods. The study was done using the original neurovascular unit model in vitro. The NVU consisted of three cell types: neurons, astrocytes, and cerebral endothelial cells derived from rats. Hypoxia was induced by incubating cells with sodium iodoacetate for 30 min at37 °C in standard culture conditions.Results. We investigated the role of connexin 43 in the regulation of cell interactions within the NVU in normal and hypoxic injury in vitro. We found that astrocytes were characterized by high levels of expression of Cx43 and low level of CD38 expression, neurons demonstrated high levels of CD38 and low levels of Cx43. In hypoxic conditions, the expression of Cx43 and CD38 in astrocytes markedly increased while CD38 expression in neurons decreased, however no changes were found in endothelial cells. Suppression of Cx43 activity resulted in down-regulation of CD38 in NVU cells, both in physiological conditions and at chemical hypoxia.Conclusion. Thus, the Cx-regulated intercellular NAD+-dependent communication and secretory phenotype of astroglial cells that are the part of the blood-brain barrier is markedly changed in hypoxia.

3. Patent number 2494670 Russian Federation. Method of correction of advanced hand activity using sensitive glove / S.V. Prokopenko, E.Yu. Mazheyko. – № 2011150588; publ. 10.10.2013, Bul. № 17. – P. 8. 4. Barker R.N., Gill T.J., Brauer S.G. Factors contributing to upper limb recovery after stroke: a survey of stroke survivors in Queensland, Australia // Disabil Rehabil. – 2007. –Vol. 29. – P. 981-989.5. BroerenJ., Rydmark M., Bjirkdahl A., Stibrant Sunnerhagen K. Assessment and Training in a 3-Dimensional Virtual Environment With Haptics: A Report on 5 Cases of Motor Rehabilitation in the Chronic Stage After Stroke // Neurorehabil Neural Repair. – 2007. – Vol. 21, № 2. – P. 180-189. 6. Fasoli S.E., Krebs H.I., Stein J., Frontera W.R., Hogan N.. Robotic therapy for chronic motor impairments after stroke: follow-up results // Arch. Phys. Med. Rehabil. – 2004. – Vol. 85. – P. 1106-1111. 7. Hammer Ann M., Lindmark B. Effects of Forced Use on Arm Function in the Subacute Phase After Stroke: A Randomized, Clinical Pilot Study // Physical Therapy

The chemical composition of the CHCl3 fraction of the total extract (95% EtOH) of Alfredia cernua (L.) Cass aerial part included triterpene alcohols (α-amyrin, β-amyrin, moretenol, lupeol) and butyrolignan (arctiin) in addition to various simple phenols and organic acids and their esters. It was established that the triterpene alcohols and arctiin demonstrated marked nootropic activity that was comparable with that of the whole CHCl3 fraction.

We studied the model to figure out the factors that may affect and retard the development of Alzheimer’s disease. The experimental rats have been kept in two kinds of environment: standard one vs. enriched one, and amiloid protein has been injected to both groups of rats to simulate Alzheimer’s disease. It is found the enriched environment is the key factor to retard the development of neurodegenerative disorder.

The peculiarities in expression of transport proteins and the proteins implicated in the control of glycolysis by the cellular components of neurovascular units were examined in animals of different age under normal conditions and after modeled perinatal stress or hypoxic brain injury. In both cases, the specialties in expression of transport proteins in ontogenesis were revealed. The perinatal hypoxic brain injury resulted in up-regulation of MCT1, MCT4, and GLUT4 expression in endotheliocytes of hippocampal microvessels accompanied by transient elevation of HIF-1α and GSK3 expression.

Neuroinflammation is as an important component of pathogenesis in many types of brain pathology. Immune mechanisms regulate neuroplasticity, memory formation, neurogenesis, behavior, brain development, cognitive functions, and brain metabolism. It is generally believed that essential homeostatic functions of astrocytes – astroglia-neuron metabolic coupling, gliovascular control, regulation of proliferation, and migration of cells in the neurogenic niches – are compromised in neuroinflammation resulting in excitotoxicity, neuronal and glial cell death, and alterations of intercellular communication. Viral neuroinfection, release of non-coding RNAs from the cells at the sites of brain injury or degeneration, and application of siRNA or RNA aptamers as therapeutic agents would require dsRNA-sensing pathways in the cells of neuronal and non-neuronal origin. In this review, we analyze the data regarding the role of astrocytes in dsRNA-initiated innate immune response in neuroinflammation and their contribution to progression of neurodegenerative and neurodevelopmental pathology.

Perinatal hypoxic-ischemic brain injury is a relevant medical and social problem. Among many pathological processes in the neonatal period perinatal hypoxic-ischemic injury is a major cause of further hemorrhage, necrotic and atrophic changes in the brain. This review presents recent data on the basic mechanisms of the hypoxic-ischemic brain injury along the concept of neurovascular unit (neurons, astrocytes, endothelial cells, pericytes) with the focus on alterations in cell-to-cell communication. Pathological changes caused by ischemia-hypoxia are considered within two phases of injury (ischemic phase and reperfusion phase). The review highlights changes in each individual component of the neurovascular unit and their interactions. Molecular targets for pharmacological improvement of intercellular communication within neurovascular unit as a therapeutic strategy in perinatal brain injury are discussed.

This review summarizes the literature and our own data regarding the role of NAD+-glycohydrolase/CD38- controlled molecular mechanisms of hypothalamic and pituitary oxytocin secretion in social behavior regulation. Current approaches to the modulation of both CD38 expression and brain cell activity that represent prospective treatments for disorders associated with altered social behavior are discussed. © 2013 Bentham Science Publishers.

The review contains current data on molecular mechanisms which control NAD+ homeostasis in brain cells. It also deals with the role of NAD+-converting enzymes in regulation of functional activity, viability and intercellular communication of neuronal and glial cells. Special attention is paid to involvement of CD38 into regulation of NAD+ levels in brain cells in normal and pathological conditions.

Antihypoxic and nootropic activities of extracts from aerial parts of wild and cultivated Alfredia cernua (L.) Cass. were studied on the models of pressure chamber hypoxia, open field test, and passive avoidance conditioning. The extracts of Alfredia cernua promoted retention of the orientation reflex and passive avoidance conditioned response and normalized orientation and exploratory activities disordered as a result of hypoxic injury. The efficiency of the extracts was superior to that of piracetam by the effect on retention of passive avoidance response throughout the greater part of the experiment. Nootropic activity of cultivated Alfredia cernua was not inferior to that of the wild plant.

It is established, that Alfredia cernua extracts possess nootropic properties. The extracts favor improvement of the indices of orientation and exploration behavior, retention of the passive avoidance reflex upon hypoxic shock, and increase in physical work capacity in mice. The most pronounced effect was observed upon the administration of a 95% ethanol extract of Alfredia cernua.