INSTITUTE OF CYTOLOGY OF THE RUSSIANACADEMY OF SCIENCES

Laboratory of Cell Physiology
Head of the Laboratory
A. A. Vereninov
PhD., DSc.
phone: (812) 297-38-02
e-mail:
The Laboratory Staff:
4 researchers (4 PhDs), 2 laboratory technicians /
The Corresponding Member of the USSR Academy of Sciences D.N. Nasonov simultaneously with the organization of the Institute of Cytology founded the Laboratory of Cell physiology. From 1957 to 1985 the Director of the Institute, Corresponding Member of the USSR Academy of Sciences, A.S. Troshin, headed the Laboratory. From 1986 until now the Laboratory is headed by A.A. Vereninov, PhD., DSc.
The Laboratory had prepared several known scientists who later became heads of other laboratories and groups of the Institute: A.A. Lev (Group of physical chemistry of cell membranes, 1976), Academician N.N. Nikolsky, Director of Institute of Cytology in 1988-2004, Corresponding Member of the USSR Academy of Sciences G.N. Mozhaeva (Laboratory of ion channels of cell membranes, 1985), S.A. Krolenko, PhD., DSc. (Laboratory of biological cytology and cytochemistry, 1985). Pioneer trends in the cell study at the Laboratory were initiated by S.V. Levin, V.V. Malev, A.L. Krol (optical studies of cell motility underlying micromovement of nerve fiber during action potential and flickering erythrocytes and other non-excitable cells), A.B. Kaulin (study of anisotropic cell organelles by fluorescence polarization), and I.I. Marakhova (study of ion transport in proliferating cell cultures).
After budding of daughter laboratories from the Laboratory of Cell physiology, two directions of work, both originating from the interests of D.N. Nasonov and A.S. Troshin, have been developed: (1) optical study of living cells and (2) study of ion balance in animal cells.
The most remarkable results in the first direction were the discovery of movement of nerve fiber during action potential and the finding that submicron local movements of cell surface within the range 0.2-30 Hz, which are observed in numerous non-muscle cells, were due not to the thermal or electrostriction fluctuations of lipid bilayer, but due to ATP-dependent motility of the membrane skeleton.
The main publications on the first direction of work:
Levin S.V., Golfand K.A. 1980. Reversible displacement of crab axon at action potential. Tsitologiya. 22 (6) : 717-721. Levin S.V., Golfand K.A. 1984. Rapid transverse movement of crab axon at action potential. Tsitologiya. 26 (8) : 920-926. Levin S.V. et al. 1986. Wave of local shortening and lengthening of single crab axon during action potential. Tsitologiya. 28 (12) : 1307-1315. Levin S.V., Malev V.V. 1987. Effect of elastic properties of crab axon sheath on the nerve fiber movement during action potential. Tsitologiya. 29 (5) : 569-575. Krol A.Yu., Grinfeldt M.G., Levin S.V., Smilgavichus A.D. 1990. Local mechanical oscillations of the cell surface within the range 0.2-30 Hz. Eur. Biophys. J. 19 : 93-99. Levin S., Korenstein R. 1991.Membrane fluctuations in erythrocytes are linked to MgATP- dependent dynamic assembly of the membrane skeleton. Biophys. J. 60 : 733-737. Krol A.Yu., Malev V.V., Grinfeldt M.G. 1993. Spectral characteristics of spontaneous oscillations in erythrocytes and their ghosts. Biol. Mem. 6 : 701-710. Rubashkin A., Iserovich P., Hernandez J.A., Fischbarg J. 2005. Epithelial fluid transport: protruding macromolecules and space charges can bring about electro-osmotic coupling at the tight junctions. J. Membrane Biol. 208 : 251-263. Tuvia S., Almagor A., Bitler A., Levin S., Korenstein R., Yedgar S. 1997. Cell membrane fluctuations are regulated by medium macroviscosity: evidence for a metabolic driving force. PNAS USA. 94 : 5045-5049. Tuvia S., Levin S., Bitler A., Korenstein R. 1998. Mechanical fluctuations of the membrane-skeleton are dependent on F-actin ATPase in human erythrocytes. J. Cell Biol. 141 : 1551-1561. Krol A.Yu., Malev V.V., Vereninov A.A. 2000. Laser dark-field microscope: investigation of the motility of submicroscopic structures. Proc. SPIE. 4064 : 45-48. Grinfeldt M.G., Krol A.Yu., Malev V.V. 2000. Correlation between the oscillatory and adhesion activities in erythroid cells. Membr. Cell Biol. 13 : 347-355. Krol A.Yu., Grinfeldt M.G., Vereninov A.A., Malev V.V. 2000. Role of actin cytoskeleton in generation of the surface oscillations of erythrocyte hosts. Biol. Membrany. 17 (1) : 60-66.
In the early 1960-ies, studies of cell ion balance were directed on reinvestigation of the two fundamental principles which could underlie asymmetrical distribution of ions between the cytoplasm and the medium: the principle of equilibrium between different phases, which had been promoted by D.N. Nasonov and A.S. Troshin as alternative to t he classical membrane theory since 1930-ies, and the principle of permanent ion pumping, which had been advanced later. The study of 24Na, 22Na and 42K exchange in frog muscle were performed and the data previously considered as an argument for presence of "free" and "bound" K+ in muscle were reinterpreted on the pump-leak basis. Results of this work were summarized in A.A. Vereninov's monograph "Ion transport across the cell membrane. Analysis of fluxes" (1978) in which all pro and contra for the "bulk phase" and "pump-leak" concepts were reviewed, the possibility of determination of individual activity coefficients of ions in the cytoplasm-phase was criticized and plenty of current evidence was presented indicating that the fluxes considered as a "leak" should be attributed to special ion-transporting systems.
Studies of ion transport in proliferating cell cultures began in the Laboratory since the late 1970-ies. It has been found that transport of Na+ and K+ changes significantly during transition of resting cells to proliferation and that high total K+ and Na+ content per g protein is a peculiar feature of all proliferating cells. In spite of the remarkable difference between resting and proliferating cells by K+, Na+ content and fluxes, the contemporary popular idea that cell malignancy was associated with specific alteration of cell ion transport was found to be false. Results of the first years of work on cell cultures are resumed in the book: A.A. Vereninov, I.I. Marakhova "Ion transport in cultured cells" (1986). Work with the long living cell cultures opened new opportunity for studying long-term physiological regulation of cell ion and water balance. Monovalent ion transport and expression of mRNAs of a group of ion transporters (АТР1В1, NHE1, NKCC1), along with several other regulators (p53, Bcl-2, hSGK) and the reference β-actin and GAPDH mRNAs, were studied in human lymphocytes activated to proliferation by mitogens. Functional, protein and transcriptional regulation of the Na+ and K+ transport were revealed at the prereplicative stage of the lymphocyte G0-G1 transition. Comparative analysis of the expression profiles for different mRNAs in lymphocytes from one donor and for one and the same mRNA in lymphocytes from different donors was performed. It was found that the node of the signaling network which is responsible for the similarity of the expression profiles for certain mRNAs ("group" response) should be located before the elements producing their inter-donor variability (Vereninov et al., 2001a, b).
For the last decade, the major pursuit in the Laboratory is the understanding of the role of monovalent ions in apoptosis. Human lymphoid cell line U937 is used as a main object, determination of caspase activity, confocal microscopy, and flow cytometry are applied for testing apoptosis; measurement of cell buoyant density is used for determination of cell shrinkage-dehydration; and radioisotope and flame emission analysis is the main method for studying cell ion balance. It has been found that no cell shrinkage occurs in case of apoptosis of U937 cells induced by etoposide in contrast to apoptosis induced by staurosporine when cell shrinkage takes place. This finding indicates that classical definition of apoptosis, as a "shrinkage-necrosis" should be revised. It has been shown that the lack of cell dehydration during etoposide induced apoptosis of U937 cells is not due to peculiarities of cells or inducer of apoptosis, but due to a certain combination of cell and inducer. Changes of the water, K+, Na+ and Cl- content have been estimated quantitatively in case of apoptosis with cell shrinkage like that in U937 cell treated with staurosporine. The fluxes of 22Na, Rb+ and 36Cl across plasma membrane of U937 cells were studied under the normal conditions and during apoptosis cause by staurosporine. It is found that the rate of equilibration of inward and outward fluxes of monovalent ions is much higher than the rate of alteration of ion and water balance associated with apoptosis. It means that the latter should be considered as a slow drift of the balanced state (Vereninov et al., 2006, 2007; Yurinskaya et al., (Юринскаяидр.,) 2010).
The contribution of the major ion pathways, i.e. the sodium pump, K+, Na+ and Cl- channels and Na-Cl symport system to the transmembrane ion turnover has been analyzed by mathematical modeling of the overall ion flux balance (Vereninov et al., 2004a, b, 2006, 2007; Rubashkin et al., 2010). Several scenarios of alteration of ion balance in U937 cells induced to apoptosis by staurosporine are revealed by matching the calculated values and experimental data obtained by measurement of cell water, K+, Na+ and Cl- content and Rb+, 22Na, Na+, and 36Cl net and unidirectional fluxes. It is found that in dependence of cell properties and the depth of apoptosis the apoptotic changes of ion and water balance in some cases are due to the sodium pump degradation in accompany with the closing of Na+ channels and reducing of Na-Cl symport, whereas in other cases changes in ion balance are due to the opening of the K+ channels without the pump degradation. The Laboratory pioneers in analysis of the balance of the monovalent ion fluxes via all major pathways across plasma membrane of proliferating and apoptotic cells.
Electro-osmosis as a driving force in paracellular fluid transport in epithelia is studied in the Laboratory by A.A. Rubashkin. The role of the fixed electric charge in tight junctions, e.g. associated with claudine has been analyzed theoretically and examined in experiments with corneal endothelium due to collaboration with colleagues from Columbia University (USA). Their paper (Rubashkin et al., 2005) was selected and reviewed by "Faculty of 1000 Biology" as one of the most interesting 1000 recent studies in biology.

Apoptosis of U937 cells
Confocal images of human lymphoid cells U937 stained with acridine orange, ethidium bromide, and the lipophilic dye RH414. a - control cells, b, c - apoptosis caused by staurosporine (b) or etoposide (c), d - apoptotic bodies (Yurinskaya et al. Cell Physiol. Biochem. 2005. 16 : 155-162).
The main publications on the second direction of work since 1995:
Marakhova I.I., Vereninov A.A., Toropova F.V., Vinogradova T.A. 1995. Long-term enhancement of Na, K-ATPase pump during blasttransformation of human lymphocytes is controlled first by translational, then by transcriptional mechanisms. FEBS Lett. 368 : 110-112. Vereninov A.A., Glushankova L.N., Rubashkin A.A. 1995. The role of ion transport in long-term cell volume regulation. Mathematical model and live lymphoid cells. Tsitologiya. 37 (12) : 1151-1166. Vereninov A.A., Glushankova L.N., Rubashkin A.A. 1997. Effect of NaK2Cl symport and chloride channel permeability on the ion flux balance and ion distribution in animal cells of different types. Tsitologiya. 39 (8) : 727-739. Marakhova I.I., Vereninov A.A., Toropova F.V., Vinogradova T.A. 1998. Na, K-ATPase pump in activated human lymphocytes: on the mechanisms of rapid and long-term increase in K influxes during blasttransformation. Biochim. Biophys. Acta. 1368 : 61-72. Marakhova I.I., Vereninov A.A., Vinogradova T.A. Toropova F.V. 1998. Cyclosporin A inhibits long-term activation of Na,K pump in phytohemagglutinin-stimulated human lymphocytes. Membr. Cell Biol. 12 : 363-374. Marakhova I.I., Ivanova A.E., Toropova F.V., Vereninov A.A., Vinogradova T.A. 1999. Functional expression of the Na/K pump is controlled via a cyclosporin A-sensitive signalling pathway in activated human lymphocytes. FEBS Letters. 456 : 285-289. Кrol А.Yu. Malev V.V., Vereninov A.A. 2000. Laser dark-field microscope: investigation of the motility of submicroscopic structures. Proc. Soc. Photo-Optical Instr. Eng. 4064 : 45-48. Krol A.Yu., Grinfeldt M.G, Vereninov A.A., Malev V.V. 2000. Role of actin cytoskeleton in generation of surface oscillations of erythrocyte hosts. Biol. Membrany. 17 (1) : 60-66. Vereninov A.A., Vasilieva I.O., Yurinskaya V.E., Matveev V.V., Glushankova L.N. 2001a. A study of the "group" expression of ATP1B1, NHE1, NKCC1, β-actin, GAPDH, p53, BCl-2, and hSGK mRNAs in human lymphocytes activated with phytohemagglutinin. Tsitologiya. 43 (6) : 602-612. Fillon S., Warntges S., Matskevitch J., Moschen I., Setiawan I., Gamper N., Feng Y.X., Stegen C., Friedrich B., Waldegger S., Broer S., Wagner C.A., Huber S.M., Klingel K., Vereninov A., Lang F. 2001. Serum- and glucocorticoid-dependent kinase, cell volume, and the regulation of epithelial transport. Comp. Biochem. Physiol., Part A. 130 : 367-376. Vereninov A.A., Vassilieva I.O., Yurinskaya V.E., Matveev V.V., Glushankova L.N., Lang, F., Matskevich J.A. 2001b. Differential transcription of ion transporters, NHE1, ATP1B1, NKCC1 in human peripheral blood lymphocytes activated to proliferation. Cell. Physiol. Biochem. 11 (1) : 19-26. Zuniga F.A., Shi G., Haller J.F., Rubashkin A., Flynn D.R., Iserovich P., Fischbarg J.2001. A three-dimensional model of the human facilitative glucose transporter Glut1. J. Biol. Chem. 276 : 44970-44975. Sanchez J.M., Li Y., Rubashkin A., Iserovich P., Wen Q., Ruberti J.W., Smith R.W., Rittenband D., Kuang K., Diecke F.P.J., Fischbarg J. 2002. Evidence for a central role for electro-osmosis in fluid transport by corneal endothelium. J. Membrane Biol. 187 : 37-50. Vereninov A.A., Volgareva E.V., Matveev V.V., MoshkovA.V., Rozanov Yu.M., Shirokova A.V., Yurinskaya V.E. 2003. Water and ion balance of rat thymocytes in apoptosis induced by dexamethasone or etoposide. Ion-osmotic model of cell volume decrease. Tsitologiya. 45 (5) : 500-509. Vereninov A.A., Goryachaya T.S., Matveev V.V., Moshkov A.V., Rozanov Yu.M., Sakuta G.A., Shirokova A.V., Yurinskaya V.E. 2004a. Cell shrinkage in apoptosis is not obligatory. Apoptosis of U937 cells induced by staurosporine and etoposide. Tsitologiya. 46 (7) : 609-619. Vereninov A.A., Yurinskaya V.E., Rubashkin A.A. 2004b. The role of potassium, potassium channels, and symporters in apoptotic cell volume decrease: Experiment and theory. Doklady Biological Sciences, 398 : 417-420. Yurinskaya V.E., Moshkov A.V., Rozanov Yu.M., Shirokova A.V., Shumilina E.V., Vassilieva I.O., Lang F., Volgareva E. V, Vereninov A.A. 2005a. Thymocyte K+, Na+ and water balance during dexamethasone and etoposide induced apoptosis. Cell. Physiol. Biochem. 16 (1-3) : 15-22. Yurinskaya V.E., Goryachaya T.S., Moshkov A.V., Rozanov Yu.M., Sakuta G.A, Shirokova A.V., Shumilina E.V., Vassilieva I.O., Lang F., Vereninov A.A. 2005b. Potassium and sodium balance in U937 cells during apoptosis with and without cell shrinkage. Cell. Physiol. Biochem. 16 : 155-162. Lang F., Foller M., Lang K., Lang P., Wieder T., Ritter M., Gulbins E., Vereninov A., Huber S. 2005. Ion channels in cell proliferation and apoptotic cell death. J. Membr. Biol. 205 (3) : 147-157. Rubashkin A., Iserovich P., Hernandez J., Fischbarg J. 2005. Epithelial fluid transport: protruding macromolecules and space charges can bring about electro-osmotic coupling at the tight junctions. J. Membr. Biol. 208 : 251-263. Fischbarg J., Diecke F.P.J., Iserovich P., Rubashkin A. 2006. The role of the tight junction in paracellular fluid transport across corneal endothelium. Electro-osmosis as a driving force. J. Membr. Biol. 210 (2) : 117-130. Rubashkin A.A.2006. A model of electro-osmosis in a leaky tight junction of epithelial cells. Doklady Biochemistry and Biophysics. 407 : 71-73. Lang F., Shumilina E., Ritter M., Gulbins E., Vereninov A., Huber S.M. 2006. Ion channels and cell volume in regulation of cell proliferation and apoptotic cell death. Contrib. Nephrol. 152 : 142-160. Vereninov A.A. Yurinskaya V.E., Rubashkin A.A. 2006. Apoptotic shrinkage of lymphoid cells: a model of changes in ion flux balance. Doklady Biochemistry and Biophysics. 411 : 356-360. Vereninov A.A., Goryachaya T.S., MoshkovA.V., Vassilieva I.O., Yurinskaya V.E., Lang F., Rubashkin A.A. 2007. Analysis of the monovalent ion fluxes in U937 cells under the balanced ion distribution: Recognition of ion transporters responsible for changes in cell ion and water balance during apoptosis. Cell Biol. Intern. 31 (4) : 382-393. Shirokova A.V. 2007. Apoptosis signaling pathways and cell ion and water balance. Cell and Tissue Biology. 1 (3): 215-224. Rubashkin A.A., Iserovich P.2007. A new approach to the selectivity of ion channels: nonlocal electrostatic consideration. Dokl. Biophys. Biochem. 417 : 302-305. Lang F., Foller M., Lang K., Lang P., Ritter M., Vereninov A., Szabo I., Huber S.M., Gulbins E. 2007. Cell volume regulatory ion channels in cell proliferation and cell death. Methods in Enzymology. Osmosensing and Osmosignaling. 428 : 209-225. Lang F., Gulbins E., Szabo I., Vereninov A.A., Huber S.M. 2008. Ion channels, cell volume, cell proliferation and apoptotic cell death. In: Sensing with Ion Channels. Springer Series in Biophysics 11. B.Martinac (ed.). Vereninov A.A., Rubashkin A.A., Goryachaya T.S., Moshkov A. V., Rozanov Y.M., Shirokova A.V., Strelkova E.G., Lang F., Yurinskaya V.E. 2008. Pump and channel K (Rb+) fluxes in apoptosis of human lymphoid cell line U937. Cell Physiol. Biochem. 22 : 187-194. A.A. Rubashkin, V.E. Yurinskaya, A.A. Vereninov. 2010. Computation of the K+, Na+ and Cl- fluxes through membrane of living cell with Na+/K+ pump, NKCC, NC cotransporters and ion channels with and without non-Goldman's rectification in K+ channels. Norma and apoptosis. Tsitologiya.52 (in press). V.E. Yurinskaya, T.S. Goryachaya, A.A. Rubashkin, A.V. Shirokova, A.A. Vereninov. 2010. Changes in K+, Na+ and Cl- balance and K+ and Cl- fluxes in U937 cells induced to apoptosis by staurosporine. On the mechanism of cell dehydration in apoptosis. Tsitologiya. 52 (in press).