State Key Laboratory of Molecular and Cellular Biology

Kiev, Ukraine

State Key Laboratory of Molecular and Cellular Biology

Kiev, Ukraine
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Fedorenko O.A.,Bogomoletz Institute of Physiology | Fedorenko O.A.,State Key Laboratory of Molecular and Cellular Biology | Popugaeva E.,Saint Petersburg State Polytechnic University | Enomoto M.,University of Toronto | And 4 more authors.
European Journal of Pharmacology | Year: 2013

The inositol-1,4,5-trisphosphate receptors (InsP3Rs) are the major intracellular Ca2+-release channels in cells. Activity of InsP3Rs is essential for elementary and global Ca2+ events in the cell. There are three InsP3Rs isoforms that are present in mammalian cells. In this review we will focus primarily on InsP3R type 1. The InsP3R1 is a predominant isoform in neurons and it is the most extensively studied isoform. Combination of biophysical and structural methods revealed key mechanisms of InsP3R function and modulation. Cell biological and biochemical studies lead to identification of a large number of InsP3R-binding proteins. InsP3Rs are involved in the regulation of numerous physiological processes, including learning and memory, proliferation, differentiation, development and cell death. Malfunction of InsP3R1 play a role in a number of neurodegenerative disorders and other disease states. InsP3Rs represent a potentially valuable drug target for treatment of these disorders and for modulating activity of neurons and other cells. Future studies will provide better understanding of physiological functions of InsP3Rs in health and disease. © 2013 Elsevier B.V. All rights reserved.


Sukhanova K.Yu.,Institute of Physiology | Sukhanova K.Yu.,State Key Laboratory of Molecular and Cellular Biology | Harhun M.I.,St George's, University of London | Bouryi V.A.,Institute of Physiology | And 4 more authors.
Pharmacological Reports | Year: 2013

Background: There is growing evidence suggesting involvement of L-type voltage-gated Ca2+ channels (VGCCs) in purinergic signaling mechanisms. However, detailed interplay between VGCCs and P2X receptors in intracellular Ca2+ mobilization is not well understood. This study examined relative contribution of the Ca2+ entry mechanisms and induced by this entry Ca2+ release from the intracellular stores engaged by activation of P2X receptors in smooth muscle cells (SMCs) from the guinea-pig small mesenteric arteries. Methods: P2X receptors were stimulated by the brief local application of αβ-meATP and changes in [Ca 2+]i were monitored in fluo-3 loaded SMCs using fast x-y confocal Ca2+ imaging. The effects of the block of L-type VGCCs and/or depletion of the intracellular Ca2+ stores on αβ-meATP- induced [Ca2+]i transients were analyzed. Results: Our analysis revealed that Ca2+ entry via L-type VGCCs is augmented by the Ca 2+-induced Ca2+ release significantly more than Ca 2+ entry via P2X receptors, even though net Ca2+ influxes provided by the two mechanisms are not significantly different. Conclusions: Thus, arterial SMCs upon P2X receptor activation employ an effective mechanism of the Ca2+ signal amplification, the major component of which is the Ca2+ release from the SR activated by Ca2+ influx via L-type VGCCs. This signaling pathway is engaged by depolarization of the myocyte membrane resulting from activation of P2X receptors, which, being Ca 2+ permeable, per se form less effective Ca2+ signaling pathway. This study, therefore, rescales potential targets for therapeutic intervention in purinergic control of vascular tone. Copyright © 2013 by Institute of Pharmacology Polish Academy of Sciences.


Storozhuk M.,Ukrainian Academy of Sciences | Storozhuk M.,State Key Laboratory of Molecular and Cellular Biology | Yatsenko N.,Ukrainian Academy of Sciences | Nikolaenko L.,Ukrainian Academy of Sciences | And 2 more authors.
Pharmacologyonline | Year: 2015

Pharmacologicalanalysisisoneofthemajortoolsinphysiology.Therefore,itisimportanttobeawareofnon-specificeffectsofreceptoragonists/antagonists.Besideotherquestions,pharmacologicalanalysisiswidelyusedtostudyaquestionofcoreleaseofdifferentneurotransmitters.Becauseconcentrationoftransmitterinsynapticcleftisratherhigh,andonlycompetitiveantagonistsareoftenavailable,highconcentrationoftheantagonistsarerequiredtoblockthereceptorofinterest,makingquestionofspecificityevenmoreimportant.Consideringimportantearlierobservation(Nakazawaetal.,1995)regardingsuppressingside-effectofbroadspectrumantagonistofP2X-receptorssuraminoncurrentsevokedbyexogenousGABAapplication(GABA-currents),westudiedeffectsoftwootherantagonistsNF279(8,8'-[Carbonylbis(imino-4,1-phenylenecarbonylimino-4,1-phenylenecarbonylimino)]bis-1,3,5-naphthalenetrisulfonicacidhexasodiumsalt)andPPADS(pyridoxalphosphate-6-azophenyl-2',4'-disulfonicacid)onGABA-currentsinisolatedhippocampalneurons.WefoundthatNF279(20μM)andPPADS(50μM)decreasedGABAcurrents:NF279–to~62%;PPADSto~71%respectively.Additionally,wehaveconfirmedearlierobservationregardingsuppressingeffectofsuramin(20μM)onGABA-currents.WeconcludethatinterpretationofdataobtainedusingrelativelyhighconcentrationsofNF279,PPADSandsuraminfordeterminingthefunctionalroleP2Xreceptorincomplexneuralnetworksshouldbecareful.OurresultsarealsoessentialforimprovingtheselectivityofP2X-receptorantagonistsversusGABA-receptors. © 2015, SILAE (Italo-Latin American Society of Ethnomedicine). All Rights Reserved.


Fedorenko Y.A.,Bogomolets Institute of Physiology of the NAS of Ukraine | Fedorenko Y.A.,State Key Laboratory of Molecular and Cellular Biology
Neurophysiology | Year: 2016

Inositol 1,4,5-trisphosphate receptors (IP3Rs) play a key role in intracellular calcium signaling. Up to the present time, the question on the existence of only one level of the unitary conductance of such receptors or a few levels of such conductance remained open. In experiments on the isolated nuclei of Purkinje cerebellar neurons of rats, we examined changes in the conductance of channels of these receptors localized on the internal membrane of the nuclear envelope, which were related to voltage variations. In all cases, these channels demonstrated only one level of the unitary conductance; no sublevels were found within a –100 mV to 100 mV range. Suppression of activity of IP3Rs at negative potentials is determined by a decrease in the probability of the open state of the channel. Thus, a hypothesis on the existence of a few levels of the IP3R channel conductance in the examined object has not been confirmed; the release of Ca2+ through channels of these receptors demonstrates a quantum nature. © 2016 Springer Science+Business Media New York


Prevarskaya N.,French Institute of Health and Medical Research | Prevarskaya N.,Lille University of Science and Technology | Ouadid-Ahidouch H.,Laboratory of Cellular and Molecular Physiology | Skryma R.,French Institute of Health and Medical Research | And 2 more authors.
Philosophical Transactions of the Royal Society B: Biological Sciences | Year: 2014

Cancer involves defects in the mechanisms underlying cell proliferation, death and migration. Calcium ions are central to these phenomena, serving as major signalling agents with spatial localization, magnitude and temporal characteristics of calcium signals ultimately determining cell's fate. Cellular Ca2+ signalling is determined by the concerted action of a molecular Ca2+handling toolkit which includes: active energy-dependent Ca2+ transporters, Ca2+-permeable ion channels, Ca2+-binding and storage proteins, Ca2+dependent effectors. In cancer, because of mutations, aberrant expression, regulation and/or subcellular targeting of Ca2+-handling/transport protein(s) normal relationships among extracellular, cytosolic, endoplasmic reticulum and mitochondrial Ca2+ concentrations or spatio-temporal patterns of Ca2+ signalling become distorted. This causes deregulation of Ca2+dependent effectors that control signalling pathways determining cell's behaviour in a way to promote pathophysiological cancer hallmarks such as enhanced proliferation, survival and invasion. Despite the progress in our understanding of Ca2+ homeostasis remodelling in cancer cells as well as in identification of the key Ca2+-transport molecules promoting certain malignant phenotypes, there is still a lot of work to be done to transform fundamental findings and concepts into new Ca2+ transport-targeting tools for cancer diagnosis and treatment. © 2014 The Author(s) Published by the Royal Society.


Fedorenko O.A.,Bogomoletz Institute of Physiology | Fedorenko O.A.,State Key Laboratory of Molecular and Cellular Biology | Marchenko S.M.,Bogomoletz Institute of Physiology | Marchenko S.M.,State Key Laboratory of Molecular and Cellular Biology
Hippocampus | Year: 2014

Rise in Ca2+ concentration in the nucleus affects gene transcription and has been implicated in neuroprotection, transcription-dependent neuronal plasticity, and pain modulation, but the mechanism of regulation of nuclear Ca2+ remains poorly understood. The nuclear envelope is a part of the endoplasmic reticulum and may be one of the sources of nuclear Ca2+. Here, we studied ion channels in the nuclear membrane of hippocampal neurons using the patch-clamp technique. We have found that the nuclear membrane of CA1 pyramidal and dentate gyrus granule (DG), but not CA3 pyramidal neurons, was enriched in functional inositol 1,4,5-trisphosphate receptors/Ca2+-release channels (IP3Rs) localized mainly in the inner nuclear membrane. A single nuclear ryanodine receptor (RyR) has been detected only in DG granule neurons. Nuclei of the hippocampal neurons also expressed a variety of spontaneously active cation and anion channels specific for each type of neuron. In particular, large-conductance ion channels selective for monovalent cations (LCC) were coexpressed with IP3Rs. These data suggest that: (1) the nuclear membranes of hippocampal neurons contain distinct sets of ion channels, which are specific for each type of neuron; (2) IP3Rs, but not RyRs are targeted to the inner nuclear membrane of CA1 pyramidal and DG granule, but they were not found in the nuclear membranes of CA3 pyramidal neurons; (3) the nuclear envelope of these neurons is specialized to release Ca2+ into the nucleoplasm which may amplify Ca2+ signals entering the nucleus from the cytoplasm or generate Ca2+ transients on its own; (4) LCC channels are an integral part the of Ca2+-releasing machinery providing a route for counterflow of K{cyrillic}+ and thereby facilitating Ca2+ movement in and out of the Ca2+ store. © 2014 Wiley Periodicals, Inc. © 2014 Wiley Periodicals, Inc.


Fedorenko O.A.,Bogomoletz Institute of Physiology | Fedorenko O.A.,State Key Laboratory of Molecular and Cellular Biology | Popugaeva E.,Saint Petersburg State Polytechnic University | Enomoto M.,University of Toronto | And 3 more authors.
European Journal of Pharmacology | Year: 2014

The inositol-1,4,5-trisphosphate receptors (InsP3Rs) are the major intracellular Ca2+-release channels in cells. Activity of InsP3Rs is essential for elementary and global Ca2+ events in the cell. There are three InsP3Rs isoforms that are present in mammalian cells. In this review we will focus primarily on InsP3R type 1. The InsP3R1 is a predominant isoform in neurons and it is the most extensively studied isoform. Combination of biophysical and structural methods revealed key mechanisms of InsP3R function and modulation. Cell biological and biochemical studies lead to identification of a large number of InsP3R-binding proteins. InsP3Rs are involved in the regulation of numerous physiological processes, including learning and memory, proliferation, differentiation, development and cell death. Malfunction of InsP3R1 play a role in a number of neurodegenerative disorders and other disease states. InsP3Rs represent a potentially valuable drug target for treatment of these disorders and for modulating activity of neurons and other cells. Future studies will provide better understanding of physiological functions of InsP3Rs in health and disease. © 2013 Elsevier B.V. All rights reserved.


Stepanyuk A.R.,Bogomoletz Institute of Physiology | Stepanyuk A.R.,State Key Laboratory of Molecular and Cellular Biology | Borisyuk A.L.,Bogomoletz Institute of Physiology | Borisyuk A.L.,State Key Laboratory of Molecular and Cellular Biology | And 2 more authors.
PLoS ONE | Year: 2011

A new method is described that accurately estimates kinetic constants, conductance and number of ion channels from macroscopic currents. The method uses both the time course and the strength of correlations between different time points of macroscopic currents and utilizes the property of semiseparability of covariance matrix for computationally efficient estimation of current likelihood and its gradient. The number of calculation steps scales linearly with the number of channel states as opposed to the cubic dependence in a previously described method. Together with the likelihood gradient evaluation, which is almost independent of the number of model parameters, the new approach allows evaluation of kinetic models with very complex topologies. We demonstrate applicability of the method to analysis of synaptic currents by estimating accurately rate constants of a 7-state model used to simulate GABAergic macroscopic currents. © 2011 Stepanyuk et al.


Prevarskaya N.,French Institute of Health and Medical Research | Skryma R.,Lille University of Science and Technology | Shuba Y.,Ukrainian Academy of Sciences | Shuba Y.,State Key Laboratory of Molecular and Cellular Biology
Expert Opinion on Therapeutic Targets | Year: 2013

Introduction: Cancer is caused by defects in the mechanisms underlying cell proliferation, death and migration. Calcium ions are central to all of these phenomena, serving as major signalling agents with the spatial localisation, magnitude and temporal characteristics of calcium signals ultimately determining cell's fate. The transformation of a normal cell into a malignant derivative is associated with a major rearrangement of Ca2+ pumps, Na/Ca exchangers and Ca2+ channels, which leads to enhanced proliferation and invasion under compromised/impaired ability to die. Areas covered: This paper examines the changes in Ca2+ signalling and the mechanisms that underlie the passage from normal to pathological cell growth and death control. Understanding these changes and identifying the molecular players involved provide new perspectives for cancer treatment. Expert opinion: Despite compelling evidence that the disruption of Ca2+ homeostasis in cancer cells leads to the promotion of certain malignant phenotypes as well as the identification of key Ca2+-transporting molecules whose altered expression and/or function underlies pathological changes, the therapeutic utilisation of these findings for cancer treatment is still at its infancy. However, the rapid development of the field warrants the development of improved molecular Ca2+ transport-targeting tools for cancer diagnosis and treatment. © 2013 Informa UK, Ltd.


Storozhuk M.,Bogomoletz Institute of Physiology | Storozhuk M.,State Key Laboratory of Molecular and Cellular Biology | Kondratskaya E.,Bogomoletz Institute of Physiology | Kondratskaya E.,State Key Laboratory of Molecular and Cellular Biology | And 3 more authors.
Molecular Brain | Year: 2016

Rapid acidification occurring during synaptic vesicle release can activate acid-sensing ion channels (ASICs) both on pre- and postsynaptic neurons. In the latter case, a fraction of postsynaptic current would be mediated by cation-selective acid-sensing ion channels. Additionally, in both cases, activation of acid-sensing ion channels could modulate synaptic strength by affecting transmitter release and/or sensitivity of postsynaptic receptors. To address potential involvement of acid-sensing ion channels in mediation/modulation of synaptic transmission at hippocampal GABAergic synapses, we studied effects of three structurally different blockers of acid-sensing ion channels on evoked postsynaptic currents using the patch-clamp technique. We found that GABAergic postsynaptic currents, recorded below their reversal potential as inward currents, are suppressed by all the employed blockers of acid-sensing ion channels. These currents were suppressed by ~ 20 % in the presence of a novel blocker 5b (1 μM) and by ~30 % in the presence of either amiloride (25 μM) or diminazene (20 μM). In the same cells the suppression of postsynaptic currents, recorded above their reversal potential as outward currents was statistically insignificant. These results imply that the effects of blockers in our experiments are at least partially postsynaptic. On the other hand, in the case of mediation of a fraction of postsynaptic current by acid-sensing ion channels, an increase of outward currents would be expected under our experimental conditions. Our analysis of a bicuculline-resistant fraction of postsynaptic currents also suggests that effects of the blockers are predominantly modulatory. In this work we present evidence for the first time that acid-sensing ion channels play a functional role at hippocampal GABAergic synapses. The suppressing effect of the blockers of acid-sensing ion channels on GABAergic transmission is due, at least partially, to a postsynaptic but (predominantly) modulatory mechanism. We hypothesize that the modulatory effect is due to functional crosstalk between ASICs and GABAA-receptors recently reported in isolated neurons, however, verification of this hypothesis is necessary. © 2016 The Author(s).

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