Alexey Vereninov

Russian Academy of Sciences | RAS · Institute of Cytology of the Russian Academy of Sciences

Recently, we have developed software that allows, using a minimum of required experimental data, to find the characteristics of ion homeostasis and a list of all unidirectional fluxes of monovalent ions through the main pathways in the cell membrane both in a balanced state and during the transient processes. Our approach has been successfully vali...

Studying the transport of monovalent ions across the cell membrane in living cells is complicated by the strong interdependence of fluxes through parallel pathways and requires therefore computational analysis of the entire electrochemical system of the cell. Current paper shows how to calculate changes in the cell water balance and ion fluxes caus...

The work provides a modern mathematical description of animal cell electrochemical system under a balanced state and during the transition caused by an increase in external osmolarity, considering all the main ionic pathways in the cell membrane: the sodium pump, K+, Na+, Cl- electroconductive channels and cotransporters NC, KC, and NKCC. The descr...

Cation-coupled chloride cotransporters play a key role in generating the Cl– electrochemical gradient on the cell membrane, which is important for regulation of many cellular processes. However, a quantitative analysis of the interplay between numerous membrane transporters and channels in maintaining cell ionic homeostasis is still undeveloped. He...

Cation-coupled chloride cotransporters play a key role in generating the Cl ⁻ electrochemical gradient on the cell membrane which is important for regulation of many cellular processes. However, the cooperation of transporters and channels of the plasma membrane in holding the ionic homeostasis of the whole cell remains poorly characterized because...

Fluxes of monovalent ions through the multiple pathways of the plasma membrane are highly interdependent, and their assessment by direct measurement is difficult or even impossible. Computation of the entire flux balance helps to identify partial flows and study the functional expression of individual transporters. Our previous computation of unidi...

Background/aims: Sodium is a key player in the fundamental cell functions. Fluorescent probes are indispensable tools for monitoring intracellular sodium levels in single living cells. Since the fluorescence of sodium-sensitive dyes in cells is significantly different from that in an aqueous solution, the fluorescence signal is calibrated in situ...

Monovalent ions, sodium in particular, are involved in fundamental cell functions, such as water balance and electric processes, intra- and intercellular signaling, cell movement, pH regulation and metabolite transport into and out of cells. Fluorescent probes are indispensable tools for monitoring intracellular sodium levels in single living cells...

Assessing the expression of channels on the cell membrane is a necessary step in studying the functioning of ion channels in living cells. We explore, first, if endogenous VRAC can be assayed using flow cytometry and a commercially available antibody against an extracellular loop of the LRRC8A, also known as SWELL1, subunit of the VRAC channel. The...

Many evidence shows that K⁺ ions are required for cell proliferation, however, changes in intracellular K⁺ concentration during transition of cells from quiescence to cycling are insufficiently studied. Here, we show using flame emission assay that a long-term increase in cell K⁺ content per g cell protein is a mandatory factor for transition of qu...

Fluorescent probes are a popular and indispensable tool for monitoring sodium concentration in living cells in situ. Calibration of fluorescent probes inside cells commonly uses ionophores to equilibrate intracellular and external ion concentrations. Here we test this calibration method using in parallel classical flame emission assay. Suspension h...

Monovalent ions are involved in a vast array of cellular processes. Their movement across the cell membrane is regulated by numerous channels and transporters. Identification of the pathways responsible for redistribution of ions and cell water in living cells is hampered by their strong interdependence. This difficulty can be overcome by computati...

The response of fluorescent ion probes to ions is affected by intracellular environment. To properly calibrate them, intracellular concentration of the measured ion must be made equal to its extracellular concentration. In the first, computational, part of this work, we show, using the example of potassium, that the two requirements for ion equilib...

The study aims to know how the apoptotic alteration of cell ionic balance follows from the quantitatively characterised time dependent decrease in the sodium pump rate constant and changes in permeability coefficients of Cl-, K+, and Na+ channels. New experimental data on changes in cell K+, Na+, Cl-, water contents, and the Na+/K+-ATPase-mediated...

The development of ion‐sensitive fluorescent probes enables the measurement of intracellular concentrations of ions (such as H ⁺ , K ⁺ , Na ⁺ , Ca ²⁺ , etc.) in a non‐invasive manner. One major complication with their use arises from their unpredictable behavior inside the cell, which makes it necessary to calibrate them in situ . Calibration impli...

A decrease in flow cytometric forward light scatter (FSC) is commonly interpreted as a sign of apoptotic cell volume decrease (AVD). However, the intensity of light scattering depends not only on the cell size but also on its other characteristics, such as hydration, which may affect the scattering in the opposite way. That makes estimation of AVD...

Monovalent ion traffic across the cell membrane occurs via various pathways. Evaluation of individual fluxes in whole cell is hampered by their strong interdependence. This difficulty can be overcome by computational analysis of the whole cell flux balance. However, the previous computational studies disregarded ion movement of the self-exchange ty...

Hypertonic media causes cells to shrink due to water loss through aquaporin channels. After acute shrinkage, cells either regulate their volume or, alternatively, undergo a number of metabolic changes which ultimately lead to cell death. In many cell types, hypertonic shrinkage is followed by apoptosis. Due to the complex 3D morphology of skeletal...

The results of this study of the ultrastructural changes in U-937 cells in the apoptotic state are largely consistent with analogous data available in the literature. However, we also obtained original data on the ultrastructural changes in cellular organelles and immunocytochemical localization and distribution of proteasomes. It was shown that, w...

The results of this work concerning ultrastructural changes of U-937 cells in a state of apoptosis are largely in consistent with the same information available in the literature. However, we have got the original data on the ultrastructural changes of cell organelles and immune localization and distribution of proteasomes. It has been demonstrated...

Lithium transport across the cell membrane is interesting in the light of general cell physiology and because of its alteration during numerous human diseases. The mechanism of Li+ transfer has been studied mainly in erythrocytes with a slow kinetics of ion exchange and therefore under the unbalanced ion distribution. Proliferating cultured cells w...

Many vital processes in animal cells depend on monovalent ion transport across the plasma membrane via specific pathways. Their operation is described by a set of nonlinear and transcendental equations that cannot be solved analytically. Previous computations had been optimized for certain cell types and included parameters whose experimental deter...

Interrelationships between monovalent ion transport, the membrane potential, and intracellular ion and water content of an animal cell are analyzed by computation of the balance of ion fluxes across the plasma membrane when the sodium pump and electroconductive channels operate in parallel with NKCC or Na-Cl (NC) symport. The behavior of the system...

Background/Aims: Osmotic cell shrinkage is a powerful trigger of suicidal cell death or apoptosis, which is paralleled and enforced by apoptotic volume decrease (AVD). Cells counteract cell shrinkage by volume regulatory increase (RVI). The present study explored the response of human U937 cells to hypertonic solution thus elucidating the relations...

Changes in intracellular water, K+ and Na+ of U937 cells incubated in hyperosmolar medium supplemented with 200 mM sucrose have been studied. Cells were stained with acrydine orange, ethydium bromide, APOPercentage dye, which marks the phosphatidyl serine distribution on the plasma membrane; and FLICA polycaspase fluorescent dye. It was found that...

Cells dying according to the apoptotic program, unlike cells dying via an unprogrammed mode, are able to avoid swelling and osmotic bursting with membrane disruption.There are indications that apoptosis is accompanied by suppression of the Na+/K+ pump and changes in the K+ and Cl− channels. It remains unclear how ion fluxes through individual ion p...

Time course of changes in intracellular water, K+ and Na+ of U937 cells incubated in hyperosmolar medium with addition of 200 mM sucrose was studied. Ouabain-sensitive and ouabain-resistant Rb+ (K+) influxes were measured during regulatory cell volume increase (RVI) and apoptotic volume decrease (AVD). Microscopy of cells stained by Acrydine orange...

The K+, Na+, and Cl− balance and K+ (Rb+) and 36Cl− fluxes in U937 cells induced to apoptosis by 0.2 or 1 μM staurosporine were studied using flame emission and radioisotope techniques. It is found that two-thirds of the total decrease in the amount of intracellular osmolytes in apoptotic cells is accounted for by monovalent ions and one-third cons...

The balance of K+, Na+, and Cl− fluxes across the cell membrane with the Na+/K+ pump, ion channels, and Na+K+2Cl− (NKCC) and Na+-Cl− (NC) cotransport was calculated to determine the mechanism of cell shrinkage in apoptosis. It is shown that all unidirectional K+, Na+, and Cl− fluxes; the ion channel permeability; and the membrane potential can be f...

The balance of K+, Na+ and Cl- fluxes through cell membrane with the Na+/K+ pump, ion channels and NKCC and NC cotransporters is considered. It is shown that all unidirectional K+, Na+ and Cl- fluxes through cell membrane, permeability coefficients of ion channels and membrane potential can be computed for balanced ion distribution between cell and...

K+, Na+ and Cl- balance and K+ (Rb+) and 36Cl fluxes in during apoptosis of U937 cells caused by 0.2 or 1μM staurosporine were studied by flame emission and radiotracer techniques. It is found that monovalent ion redistribution accounts for 2/3 of all decrease in the amount of intracellular osmolytes in apoptotic cells while 1/3 is due to the loss...

Ouabain-sensitive (OS) and -resistant (OR) Rb(+) influx was examined in three sublines of U937 cells to compare alterations of K(+) channel permeability and the Na(+),K(+)-ATPase pump leading to the shift in ion and water balance during apoptosis induced by 0.2 and 1microM staurosporine (STS) for 4-5 h. Cell K(+), Rb(+), Na(+) and Cl(-) content was...

At some stage cell proliferation requires an increase in cell volume and a typical hallmark of apoptotic cell death is cell shrinkage. The respective alterations of cell volume are accomplished by altered regulation of ion transport including ion channels. Thus, cell proliferation and apoptosis are both paralleled by altered activity of ion channel...

Unidirectional (22)Na, Li(+) and Rb(+) fluxes and net fluxes of Na(+) and K(+) were measured in U937 human leukemic cells before and after induction of apoptosis by staurosporine (1 microM, 4 h) to answer the question which ion transporter(s) are responsible for changes in cell ion and water balance at apoptosis. The original version of the mathema...

Alterations of cell volume are key events during both cell proliferation and apoptotic cell death. Cell proliferation eventually requires an increase of cell volume, and apoptosis is typically paralleled by cell shrinkage. Alterations of cell volume require the participation of ion transport across the cell membrane, including appropriate activity...

Cell proliferation must be accompanied by increase of cell volume and apoptosis is typically paralleled by cell shrinkage. Moreover, profound osmotic cell shrinkage may trigger apoptosis. In isotonic environment cell volume changes require the respective alterations of transport across the cell membrane. Cell proliferation is typically paralleled b...

Cell proliferation and apoptosis are paralleled by altered regulation of ion channels that play an active part in the signaling of those fundamental cellular mechanisms. Cell proliferation must--at some time point--increase cell volume and apoptosis is typically paralleled by cell shrinkage. Cell volume changes require the participation of ion tran...

During the winter pre-spawning migration, lampreys Lampetra fluviatilis stop feeding, and their liver metabolism is reduced substantially. Aerobic ATP production in hepatocytes decreased to one third and ATP content decreased by 50% as compared with the values in autumn. In spite of the decrease of endogenous and phosphorylating (oligomycin-sensiti...

Staurosporine (STS) and etoposide (Eto) induced apoptosis of the human histiocytic lymphoma cells U937 were studied to determine the role of monovalent ions in apoptotic cell shrinkage. Cell shrinkage, defined as cell dehydration, was assayed by measurement of buoyant density of cells in continuous Percoll gradient. The K+ and Na+ content in cells...

The mechanism of apoptotic cell volume decrease was studied in rat thymocytes treated with dexamethasone (Dex) or etoposide (Eto). Cell shrinkage, i.e. dehydration, was quantified by using buoyant density of the thymocytes in a continuous Percoll gradient. The K+ and Na+ content of cells from different density fractions were assayed by flame emissi...

Aqueous and ionic balance in cells was studied using the cell floating density measurement in Percoll gradient for cellular water determination and flame-emission analysis for K+ and other cations determination. The experiments were carried out on the known apoptosis models - rat thymocytes, in which apoptosis was induced by dexamethasone (1 mcM, 4...

A study was made of apoptotic cell shrinkage, which is generally believed to be a hallmark of apoptosis. The two conventional models of apoptosis were used for examination of changes in cell water balance--one is apoptosis caused in human lymphoma cell line U937 by staurosporine, and the other by etoposide. Intracellular water was determined by mea...

Cell ion and water balance was studied with respect to analysis of the osmotic model of apoptotic volume decrease (AVD) in rat thymocytes under dexamethasone (1 microM, 4-6 h) or etoposide (50 microM, 5 h) treatment. Intracellular water content was determined by measurement of cell buoyant density in continuous Percoll gradient, while intracellular...

Ample pharmacological evidence points to a role of kinases in the regulation of cell volume. Given the limited selectivity of most inhibitors, however, the specific molecules involved have remained largely elusive. The search for cell volume regulated genes in liver HepG2 cells led to the discovery of the human serum- and glucocorticoid-dependent s...

Previously, we found no segregation in F2 obtained from crosses between two Dileptus anser clones differing (under the same culture conditions) in their serotypes, i.e. in their immobilization antigens (i-antigens); indeed, all the F2 clones had mixed, i.e. hybrid serotype, being immobilized simultaneously with both immune sera developed against ei...

This work, using RT PCR, studied expression of mRNAs encoding ion transporters, the Na/H antiporter (NHE1), the beta subunit of the Na,K-ATPase pump (ATP1B1), the NaK2Cl symporter (NKCC1), and some proteins unrelated to ion transport: the serum and glucocorticoid dependent kinase (hSGK), beta-actin, a glycolytic enzyme (GAPDH), and regulators of pr...

Changes in the levels of mRNAs encoding ion transporters (ATP1B1, NHE1, NKCC1), β-actin, GAPDH, regulators of proliferation and apoptosis (p53, Bcl-2) and kinase hSGK, involved in cell water regulation, were studied using RT PCR in the peripheral human lymphocytes activated with phytohemagglutinin for 4-24 h. The common, «grouped», effect that was...

We have studied the effects of three compounds on surface oscillations of human red blood cell ghosts: the P-ATPase inhibitor, suramin; the fluorescent dye of a similar structure, 1,8-anilinonaphthalene sulfonate (ANS); and subfragment 1 of skeletal muscle myosin (S1). It has been found that suramin (10 microM), ANS (100 microM) and S1 (2 mg/ml) su...

Some years ago we have built a dark-field microscope based on a catadioptric objective used as a dark-field condenser. The apparatus allowed to measure small (tens of nm) transverse displacements of light-scattering micro-objects, such as cell margins or intracellular particles. The principle of apparatus was the registration of the change in the i...

An immunosuppressant cyclosporin A (CsA) inhibits T-cell proliferation by blocking the nuclear factor of activated T-cells (NFAT) required for expression of the interleukin-2 (IL-2) gene. This work has demonstrated for the first time that in human blood lymphocytes (HBLs) activated by phytohemagglutinin (PHA), CsA at anti-proliferative doses inhibi...

The effect of the immunosuppressive drug cyclosporin A (CsA) on the K (Rb) influx, intracellular K and Na contents, and on the major parameters of lymphocyte activation have been investigated in human peripheral blood lymphocytes activated by phytohemagglutinin (PHA). CsA suppressed protein, RNA, DNA syntheses and cell proliferation by 49.8 +/- 4.3...

Functional expression of Na, K-ATPase pump as determined by ouabain-sensitive Rb influxes has been investigated in human peripheral blood lymphocytes, activated by phytohemagglutinin (PHA) from resting state to proliferation. It is found that a rapid twofold elevation of ouabain-sensitive Rb influx in response to PHA is followed by a long-term incr...

The relationships between monovalent ion fluxes via major cell membrane pathways (Na/K pump, NaK2Cl symporter, electroconductive sodium, potassium and chloride channels) and steady state transmembrane ion distribution, membrane potential and cell water content were calculated for the high potassium animal cells with high and low membrane potential....

The transition of phytohemagglutinin-activated human lymphocytes from resting state to proliferation is accompanied by a long-term increase in ouabain-sensitive Rb(K) influx which is closely related to a cyclosporin A-sensitive step of G0/G1/S progression. At least two distinct phases of the up-regulation of cation pump has been revealed: the initi...

Ion and water balance as well as flux balance in the animal cell, equilibrated to anisosmotic solutions, are computed for a cell model supplied by Na,K-ATPase pump, electroconductive ion channels and symporters Na-Cl, K-Cl and NaK2Cl. It is shown how the potency of each of the principal transporters to regulate cell volume depends on such condition...

The effects of direct influence of gravity on a cell have been studied. Potassium transport from infusoria. Tetrahymena pyriformis GL induced by the high gravitational levels which immobilize the cell (80, 140, 260 g) was examined. A minimum acceleration level at which the cells become motionless is 80 g. Kinetics of potassium output can be describ...

Increase in Na+, K(+)-ATPase mRNAs was detected in activated lymphocytes by the RT-PCR method. alpha 1 subunit mRNA gradually increased with time and by 36 h was 2.4 times higher than at the start. Increase in the beta 1 mRNA was transient reaching a maximum in the 8 h probe and declining to the initial level in the 24 and 36 h probes. The elevatio...

The relations between intracellular potassium, sodium water content and resting potential on the one hand and the ion transport parameters and intracellular electrical charge on the other hand were computed for a model of animal cell with a several ion transporters and variable intracellular charge. The case of the balanced ion distribution is cons...

Potassium (rubidium) influx, sodium and potassium contents, as well as size distribution, DNA and protein contents and synthesis have been examined in PHA-activated human lymphocytes within 0.5-72 h. A complex set of ionic events was found to include at least two stages of the increase in potassium and sodium contents per g cell protein and in ouab...

Altogether 34 patients with the nephrotic syndrome (NS) of chronic glomerulonephritis (CGN) were studied for lithium clearance (CLi). Eleven patients with edemas were enrolled in group I, 18 subjects without pronounced edemas, in group II. Out of them 7 patients suffered from membrane proliferative glomerulonephritis (group III), 7, from mesangiopr...

Tetrahymena pyriformis are known to display different physiological and biochemical properties at different steps of their growth in culture. No data available in literature on the changes of the main cations and water in the course of ciliate culture growth, although such evidence has been provided for metazoan cell culture (Vereninov, Marakhova,...

The design and operation of an automatic system for the processing of shadowgraph and interference images are described. The system includes a two-coordinate processing table with an optical system for the projection of transparent images onto the photodetector plane, an array-type photodetector, an image filter in the photodetector field, and a de...

The action of heavy water and ouabain on sodium effluxes and rubidium influxes has been measured and compared in frog muscles (m. sartorius, R. temporaria). Approximately half of muscle sodium was substituted by lithium by preliminary incubation in mixed sodium-lithium media. The ratio of the ouabain-sensitive parts of rubidium influx and sodium ef...

The sodium efflux from the frog sartorius muscle into the media of different ion composition, prepared with ordinary and heavy water, was measured by radiotracer and flame-emission techniques. About the half of the sodium in muscles was substituted for lithium. The ouabain-sensitive, as well as external potassium- and external sodium-dependent comp...

Cation transport has been investigated in three lines of Chinese ovary cells CHO-K1 during the cell culture growth. With the increase in the cell density potassium and sodium contents decreased from 1.2 to 0.8-0.5 and from 0.5 to 0.15-0.1 mmole/g protein, respectively. The time courses of potassium and sodium changes were different, and the increas...

Membrane potential and 42K, 22Na, 86Rb fluxes were measured in frog muscles immersed in sucrose--sulfate media with cation of one type. For elimination of the internal sodium and chloride ions muscles were preincubated in sucrose solution. It was found that dependence of membrane potential on the cation concentration deviated from the Goldman--Hodg...

Concentration-emission relations are presented for assayed alkaline cations within 1-100 micro M and interfering alkaline cations at concentrations 1-150 mM. All the data were obtained with the Perkin-Elmer AA spectrophotometer (AA 306) and with an air-propane flame. The advantages of flame-emission technique for measurements of cation fluxes acros...