[Show abstract][Hide abstract] ABSTRACT: N-methyl-d-aspartate (NMDA) receptors are ligand-gated nonselective cation channels mediating fast neuronal transmission and long-term potentiation in the central nervous system. These channels have a 10-fold higher permeability for Ca(2+) compared with Na(+) or K(+) and binding of the agonists (glutamate, homocysteine, homocysteic acid, NMDA) triggers Ca(2+) uptake. The present study demonstrates the presence of NMDA receptors in rat erythrocytes. The receptors are most abundant in both erythroid precursor cells and immature red blood cells, reticulocytes. Treatment of erythrocytes with NMDA receptor agonists leads to a rapid increase in intracellular Ca(2+) resulting in a transient shrinkage via Gardos channel activation. Additionally, the exposure of erythrocytes to NMDA receptor agonists causes activation of the nitric oxide (NO) synthase facilitating either NO production in l-arginine-containing medium or superoxide anion (O(2)(.-)) generation in the absence of l-arginine. Conversely, treatment with an NMDA receptor antagonist MK-80, or the removal of Ca(2+) from the incubation medium causes suppression of Ca(2+) accumulation and prevents attendant changes in cell volume and NO/O(2)(.-) production. These results suggest that the NMDA receptor activity in circulating erythrocytes is regulated by the plasma concentrations of homocysteine and homocysteic acid. Moreover, receptor hyperactivation may contribute to an increased incidence of thrombosis during hyperhomocysteinemia.
[Show abstract][Hide abstract] ABSTRACT: There is increasing evidence showing that the interplay between neuronal and immune systems may be regulated by neuromediators. However, little is known about the involvement of glutamatergic system in such neuro-immune relations. In the present study, we have shown that some intact lymphocytes express N-methyl-D: -aspartate activated receptors (NMDA receptors), an important constituent of glutamatergic system. The activation of lymphocytes with phytohemagglutinin (PHA) induces a time-dependent increase in the amount of NMDA receptor presenting cells, and NMDA stimulates this process. Immune response of such lymphocytes is suppressed and the amount of cells producing interferon gamma (IFN-gamma) in vitro is decreased to the level corresponding to intact (non-activated) cells. Furthermore, lymphocytes in the region of inflammation, induced by spinal cord injury (SCI), are also NMDA-positive. We suggest that expression of NMDA receptors in lymphocytes is regulated by central nervous system, which controls the inflammation process.
[Show abstract][Hide abstract] ABSTRACT: Using SK-N-AS human neuroblastoma cells, which co-express the alpha1 and alpha3 isoforms of the sodium pump alpha subunit, we selectively silenced either the alpha1 or alpha3 subunit by means of transfection with small interfering RNA, and investigated cell survival and the cellular response to ouabain. We found that both of the alpha subunits are essential for cell survival, indicating that substitution of one subunit for the other is not sufficient. In the presence of both alpha subunits, ouabain causes sustained activation of extracellular signal-regulated kinases 1 and 2 (Erk1/2). This activation is not affected when the alpha1 subunit is silenced. However, when alpha3 expression is silenced, ouabain-induced activation of Erk1/2 does not occur, even at a high concentration of ouabain (1 microM). Thus, ouabain-induced Erk1/2 activation is mediated in SK-N-AS cells by alpha3 only, and alpha1 does not participate in this event. This is a clear demonstration of selective involvement of a specific sodium pump alpha subunit isoform in ouabain-induced signaling.
[Show abstract][Hide abstract] ABSTRACT: Inhibition of rat neuronal Na(+)/K(+)-ATPase alpha3 isoform at low (100 nM) ouabain concentration led to activation of MAP kinase cascade via PKC and PIP(3) kinase. In contrast to ouabain-sensitive alpha3 isoform of Na(+)/K(+)-ATPase, an ouabain-resistant alpha1 isoform (inhibition with 1 mM of ouabain) of Na(+)/K(+)-ATPase regulates MAP kinase via Src kinase dependent reactions. Using of Annexin V-FITC apoptotic test to determine the cells with early apoptotic features allows to conclude that alpha3 isoform stimulates and alpha1 suppresses apoptotic process in cerebellum neurons. These data are the first demonstration showing participation of ouabain-resistant (alpha1) and ouabain-sensitive (alpha3) Na(+)/K(+)-ATPase isoforms in diverse signaling pathways in neuronal cells.
Cell Biochemistry and Function 03/2010; 28(2):135-41. DOI:10.1002/cbf.1632 · 2.13 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In our investigation, we describe the complex model of brain oxidative stress consisted of combination of experimental brain ischemia and energy metabolism violation induced by irreversible inhibitor of mitochondrial succi-nate dehydrogenase, 3-nitropropionate (3-NPA). 3-NPA causes selective degeneration of striatum neurons, which is extremely sensitive to energy deficit. This complex model allows revealing not only biochemical but also neurological symptoms in experimental animals that permits proper estimation of protective effect of different drugs on animal status. Combination of global ischemia induced by 3-vessel occlusion of major arteries supplys rat brain and subsequent 5-day reperfusion with intraperitoneal injection of 3-nitropropionic acid induces vigorous oxidative stress in brain tissues accompanied by evident neurological symptoms in Wistar rats. Such a combination of damaging factors may be considered as a new complex experimental model of brain oxidative stress permitting the evaluation of neuroprotective effect of potential therapeutic agents. Using this model, protective effect of neuropeptide carnosine was demonstrated which is in agreement with previous data.
International Journal of Clinical and Experimental Medicine 01/2010; 3(2):144-51. · 1.42 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Effects of N-methyl-D-aspartate (NMDA), aluminum and amyloid-β (Aβ) on mouse cerebellar granule cells were studied. In the presence of all three
compounds, reactive oxygen species levels and cell necrosis were increased dramatically. Mg2+ ions and D-AP5, which are known to prevent ligand binding to NMDA-activated glutamate receptors, were effective in attenuating
the neurotoxic effect induced by the presence of all three compounds. All substances tested induced activation of p42/44 MAPK
(mitogen activated protein kinase) with no cumulative effects between them. We conclude that neurotoxicity induced by aluminum
and Aβ appears at outer cell membranes and NMDA receptors take part in this process. Increase in excitotoxic effect of glutamate
in the presence of aluminum and Aβ is suggested to be a factor which provokes Alzheimer’s disease in brain neurons.
Biochemistry (Moscow) Supplement Series A Membrane and Cell Biology 12/2009; 3(4):425-430. DOI:10.1134/S1990747809040096
[Show abstract][Hide abstract] ABSTRACT: Carnosine is a neuroprotective dipeptide consisting of beta-alanine and L-histidine. It demonstrates a number of useful features, including stimulation of brain and muscle microcirculation and a rejuvenating effect on cultured cells. Its activity is based on its antioxidant and antiglycating action that, in addition to heavy metal chelation and pH-buffering ability, makes carnosine an essential factor for preventing neurodegeneration and accumulation of senile features. Recently, carnosine was successfully used to treat patients after brain stroke or patients with Parkinson disease. We conclude that carnosine can be recommended for patients under oxidative stress as a natural remedy having high efficiency and no side effects.
Rejuvenation Research 12/2009; 13(2-3):156-8. DOI:10.1089/rej.2009.0923 · 3.93 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Two novel derivatives of carnosine--(S)-trolox-L-carnosine (STC) and (R)-trolox-L-carnosine (RTC) are characterized in terms of their antioxidant and membrane-stabilizing activities as well as their resistance to serum carnosinase. STC and RTC were synthesized by N-acylation of L-carnosine with (S)- and (R)-trolox, respectively. STC and RTC were found to react more efficiently with 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) and protect serum lipoproteins from Fe(2+)-induced oxidation more successfully than carnosine and trolox. At the same time, STC, RTC and trolox suppressed oxidative hemolysis of red blood cells (RBC) less efficiently than carnosine taken in the same concentration. When oxidative stress was induced in suspension of cerebellum granule cells by their incubation with N-methyl-D-aspartate (NMDA), or hydrogen peroxide (H(2)O(2)), both STC and RTC more efficiently decreased accumulation of reactive oxygen species (ROS) than carnosine and trolox. Both STC and RTC were resistant toward hydrolytic degradation by human serum carnosinase. STC and RTC were concluded to demonstrate higher antioxidant capacity and better ability to prevent cerebellar neurons from ROS accumulation than their precursors, carnosine and trolox.
[Show abstract][Hide abstract] ABSTRACT: Hyperhomocysteinemia is a risk factor for a number of cardiovascular and neurodegenerative processes as well as a complicating factor in normal pregnancy. Toxic effects of homocysteine and the product of its spontaneous oxidation, homocysteic acid, are based on their ability to activate NMDA receptors, increasing intracellular levels of ionized calcium and reactive oxygen species. Even a short-term exposure of cells to homocysteic acid at concentrations characteristic of hyperhomocysteinemia induces their apoptotic transformation. The discovery of NMDA receptors both in neuronal tissue and in several other tissues and organs (including immunocompetent cells) makes them a target for toxic action of homocysteine. The neuropeptide carnosine was found to protect the organism from homocysteine toxicity. Treatment of pregnant rats with carnosine under conditions of alimentary hyperhomocysteinemia increases viability and functional activity of their progeny.
[Show abstract][Hide abstract] ABSTRACT: The validity of the free radical theory of aging has been recently questioned. Our aim was to test whether there is oxidative stress in tissues critically involved in accelerated aging (senescence-accelerated mice, SAM) and whether this correlates with lower glucose consumption in vivo and behavioural tests. Positron emission tomography shows that brains of old SAM-prone animals consume less glucose than young ones. Behavioural characteristics, mitochondrial peroxide production, and damage in both the central nervous system and bone marrow stem cells also indicate that SAM-prone animals age faster than SAM-resistant ones. Our results support the role of the free radical theory of aging in critical tissues involved in aging and that this correlates with glucose consumption.
[Show abstract][Hide abstract] ABSTRACT: Carnosine administration (in a daily dose of 2 g) as an additional therapy for patients with chronic discirculatory encephalopathy
(DE) results in an increase in stability of blood plasma lipoproteins toward Fe2+-induced oxidation and stabilization of red blood cells against acidic hemolysis as well as intensification of respiratory
burst of periphery blood leucocytes. At the same time, latent period of brain induced potentials, P300, decreases and the
proportion of the high-amplitude induced potentials increases. The data presented demonstrate that carnosine enhances the
efficiency of basal therapy of DE patients.
Biochemistry (Moscow) Supplement Series A Membrane and Cell Biology 03/2009; 3(1):62-65. DOI:10.1134/S1990747809010085
[Show abstract][Hide abstract] ABSTRACT: Homocysteine is a risk factor of many neurodegenerative and cardiovascular diseases. The target for its effect was suggested
to be brain neurons. Recently, it was found that homocysteine can affect immune competent cells. In this work we have shown
that, in neutrophils, homocysteine stimulates the amplitude and initial rate of respiratory burst induced by fMLP. Adenosine
receptors of the A1 and A2 types are involved in this process but not the A3 type. Phospholipase C is also involved in the
stimulation of the fMLP-induced respiratory burst by homocysteine.
[Show abstract][Hide abstract] ABSTRACT: The addition of the neuropeptide carnosine (beta-alanyl-L-histidine) as a food additive to the basic protocol of Parkinson's disease treatment results in significant improvement of neurological symptoms, along with increase in red blood cell Cu/Zn-SOD and decrease in blood plasma protein carbonyls and lipid hydroperoxides, with no noticeable change in platelets MAO B activity. The combination of carnosine with basic therapy may be a useful way to increase efficiency of PD treatment.
Rejuvenation Research 09/2008; 11(4):821-7. DOI:10.1089/rej.2008.0716 · 3.93 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Prenatal hyperhomocysteinemia induced in rats by overloading of dietary methionine (1 g/kg body mass daily) results in systemic
disordering in progeny related to an increase in the excitotoxic feature of NMDA-receptors in cerebellar neurons and memory
suppression. Administration of carnosine (100 mg/kg body mass daily) in the diet of pregnant rats with hyperhomocysteinemia
prevents both cognitive function in pups and protects cerebellar neurons from oxidative stress. The effect of carnosine is
accompanied by with the restoration of superoxide dismutase in rat brain, which is decreased during hyperhomocysteinemia from
2.07 units (control) to 1.54 units.
[Show abstract][Hide abstract] ABSTRACT: Known experimental models of Parkinson's Disease (PD) are limited by nonsimultaneous expression of physiological and biochemical features. We described a novel PD model consisting of N-methyl,4-phenyl-tetrahydropyridine (MPTP) treatment of Senescence Accelerated Mice (SAM) characteristic of increased level of reactive oxygen species aggravated by deficiency of antioxidant defense system. MPTP treatment was found to suppress locomotion and enhance the nonmotivated behavior (grooming); short-term tremor and apparent rigidity were also noted. MPTP effect was accompanied with increased level of protein carbonyls and lipid hydroperoxides indicating numerous disorders in antioxidant defense system. The brain of MPTP treated animals demonstrated higher MAO B activity and low level of SOD. Brain of control animals treated with MPTP was demonstrated by unchanged MAO B and two times decreased SOD activity; behavioral alterations in these mice being less manifested than that of SAM animals. The data presented showed that MPTP treated SAM animals are perspective model for Parkinson's disease study.
Acknowledgements: The work is supported by RFBR (## 99-04-49420; 00-04-48767).
Journal of Neurochemistry 06/2008; 81(s1):60-63. DOI:10.1046/j.1471-4159.81.s1.20_7.x · 4.24 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Three-hour incubation of rat cerebellar granule cells with 0.1 μM ouabain increases intracellular levels of Ca2+ ions and reactive oxygen species (ROS) resulting in pronounced activation of Mitogen-Activated Protein Kinase (MAPK). Higher
concentrations of ouabain induce further increases in MAPK activity. The activating effect of ouabain is attenuated by the
NMDA-receptor antagonists MK-801 and D-AP5. The data obtained suggest that similar to NMDA receptors ouabain-sensitive and
ouabain-resistant isoforms of Na+,K+-ATPase are actively involved in intracellular signaling cascades controlling proliferative activity of neuronal cells.
Biochemistry (Moscow) Supplement Series A Membrane and Cell Biology 06/2008; 2(2):156-160. DOI:10.1134/S1990747808020104
[Show abstract][Hide abstract] ABSTRACT: We evaluated possible therapeutic effect of multipotent mesenchymal stromal cells from human adipose tissue differentiated to neuronal phenotype with retinoic acid on Wistar rats subjected to toxic effect of 3-nitropropionic acid. Transplantation of mesenchymal stromal cells from human adipose tissue considerably decreased neurological symptoms, normalized exploratory activity (open field test) and long-term memory (Morris test), which correlated with normalization of pathomorphological manifestations in the brain. Destructive changes in the caudate nucleus caused by treatment with 3-nitropropionic acid (reduced size of neurons, changes in their shape, and cell edema) tended to decrease under the effect of multipotent mesenchymal stromal cells: the area of neurons increased 2-fold, the cells acquired typical round shape, cell edema decreased.
Bulletin of Experimental Biology and Medicine 05/2008; 145(4):514-9. DOI:10.1007/s10517-008-0131-5 · 0.37 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The cardioprotective effect of cardioplegic solution based on histidine-containing dipeptides was evaluated on isolated rat heart under conditions of hyporthermia and long ischemia. The use of natural dipeptides in cardioplegic solutions promoted an increase in the buffer capacity of myocardial cells and creation of an additional anti-ischemic effect under conditions of long ischemia and hypothermia.
Bulletin of Experimental Biology and Medicine 04/2008; 145(3):323-7. DOI:10.1007/s10517-008-0081-y · 0.37 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In this work, we demonstrate that homocysteic acid provokes oxidative stress in erythrocytes and decreases their hemolytic resistance, whereas the natural antioxidant carnosine  protects erythrocytes from its toxic effect. The discovered property can be used for development of new methods for protecting erythrocytes in hyperhomocysteinemia. Homocysteine and homocysteic acid (HCA), the product of spontaneous homocysteine oxidation, are important risk factors for neurodegenerative and cardiovascular diseases . These compounds induce oxidative stress in brain neurons [2, 3] and lymphocytes , resulting from their toxic effect on the nervous and immune systems. The prooxidant effect of homocysteine and homocysteic acid on cell structures can be realized via both the glutamate receptors [3, 4] and the activation of NO synthase or inhibition of Na/KATPase, as was demonstrated for the vascular endothelium [5, 6]. It is known that erythrocytes are capable of accumulating homocysteine and excreting it into the extracellular medium [7, 8]. Erythrocytes constitute the main part of blood cells; correspondingly, the toxic effect of homocysteine and HCA may cover these cells as well. However, the effect of these compounds on the erythrocyte stability has not been studied. The goal of this work was to analyze the effect of HCA on human erythrocytes and to assess the effect of carnosine on the oxidative stress in erythrocytes caused by HCA. Various factors can cause erythrocyte hemolysis, including a decreased ambient osmotic pressure, decreased pH, and oxidants [10‐12]. The resistance of erythrocytes to hemolytic action is an integral parameter characterizing their integrity and viability as well as a criterion of their physiologically native state. Two models of hemolysis were used in this work, namely, the osmotic hemolysis and the hemolysis caused by hydrochloric acid. The former takes place with a decrease in the tonicity of medium produced by diluting cell suspension with distilled water. In this process, cells swell with subsequent disruption of the cell membrane. The latter is induced by supplementing cell suspension with hydrochloric acid, which leads to a decrease in the pH in the cytoplasm, impairments in the cytoskeleton structure, and, eventually, cell swelling and destruction .
[Show abstract][Hide abstract] ABSTRACT: Activity of Na/K-ATPase isolated from bovine brain and kidney decreases during incubation with hydrogen peroxide proportionally
to the incubation time and the concentration of the oxidant. Activity suppression is accompanied by a proportional decrease
in the level of free SH-groups. Incubation of the oxidized enzyme with dithiothreitol restores the enzyme activity. Na-conformation
of the enzyme is more resistant to oxidation, whereas its conversion to the K-form increases its sensitivity to H2O2. Involvement of different conformational states of the enzyme in intracellular signaling, which occurs with the participation
of active forms of oxygen, is discussed.