Papers by Alexander Mongin
![Research paper thumbnail of [The coupling of Na,K-ATPase with the sodium channels in the plasma membranes of the brain synaptosomes of rats]](https://a.academia-assets.com/images/blank-paper.jpg)
Tsitologiia
Effect of neurotoxins veratrine (100 micrograms/ml) and tetrodotoxin (1 microM) on the binding of... more Effect of neurotoxins veratrine (100 micrograms/ml) and tetrodotoxin (1 microM) on the binding of 3H-ouabain (10(-8) M) with Na,K-ATPase of intact synaptosomes and isolated synaptic membranes was studied. The persistent opening of sodium channels in synaptosomes by veratrine results in an increase of specific binding of the labeled ligand by 20%. A similar effect was caused by Na/H exchanger monensin. Destruction of microtubules with vinblastine and colchicine has no influence on veratrine action, while depolymerization of microfilaments with cytochalasin B reverses the neurotoxin effect. In isolated synaptic membranes veratrine and tetrodotoxin stimulate ouabain binding, the absolute veratrine-induced increment being several times higher in the presence of ATP than in its absence. Since the closed vesicles of any type are not permeable to ATP and ouabain, it means that in the isolated membranes an interaction between sodium channels and Na,K-ATPase molecules takes place. In intact nerve endings such a mechanism may be operative along with the known ways of control of sodium pump and its ouabain-binding site.
![Research paper thumbnail of [3H]Taurine and D-[3H]aspartate release from astrocyte cultures are differently regulated by tyrosine kinases](https://attachments.academia-assets.com/39162682/thumbnails/1.jpg)
The American journal of physiology
H]aspartate release from astrocyte cultures are differently regulated by tyrosine kinases. Am. J.... more H]aspartate release from astrocyte cultures are differently regulated by tyrosine kinases. Am. J. Physiol. 276 (Cell Physiol. 45): C1226-C1230, 1999.-Volume-dependent anion channels permeable for Cl Ϫ and amino acids are thought to play an important role in the homeostasis of cell volume. Astrocytes are the main cell type in the mammalian brain showing volume perturbations under physiological and pathophysiological conditions. We investigated the involvement of tyrosine phosphorylation in hyposmotic medium-induced [ 3 H]taurine and D-[ 3 H]aspartate release from primary astrocyte cultures. The tyrosine kinase inhibitors tyrphostin 23 and tyrphostin A51 partially suppressed the volume-dependent release of [ 3 H]taurine in a dose-dependent manner with half-maximal effects at ϳ40 and 1 µM, respectively. In contrast, the release of D-[ 3 H]aspartate was not significantly affected by these agents in the same concentration range. The inactive analog tyrphostin 1 had no significant effect on the release of both amino acids. The data obtained suggest the existence of at least two volume-dependent anion channels permeable to amino acids in astrocyte cultures. One of these channels is permeable to taurine and is under the control of tyrosine kinase(s). The other is permeable to both taurine and aspartate, but its volume-dependent regulation does not require tyrosine phosphorylation. regulatory volume decrease; swelling-activated anion channels; excitatory amino acids; tyrosine phosphorylation; tyrphostins 0363-6143/99 $5.00
![Research paper thumbnail of [Osmotic regulation of the sodium pump in rat brain synaptosomes]](https://attachments.academia-assets.com/42404143/thumbnails/1.jpg)
Biofizika
The effect of hypoosmolality of incubation medium on the rat of ouabain-sensitive 86Rb+ transport... more The effect of hypoosmolality of incubation medium on the rat of ouabain-sensitive 86Rb+ transport in rat brain synaptosomes was studied. A decreased osmolality from 310 to 250 mOsm increased the rate of 86Rb+ uptake from 3.72 to 6.23 nmol/mg of protein min. To evaluate the involvement of cytoplasmic sodium in sodium pump stimulation inhibitors of ion channels and transport pathways able to increase [Na+]in were used. Tetrodotoxin (1 ~M), amiloride (0.5 mM) and verapamil (0.1 mM) had no influence on the osmotic response of the sodium pump. The decrease of sodium concentration in incubation medium to 15 mM, leading to a practical loss of its transmembrane gradient, did not abolish stimulation of pump. No increase in 22Na ÷ influx or intrasynaptosomal sodium content was registered at hypotonic conditions. It is suggested that osmotic regulation of Na+,K ÷-ATPase is not connected with an increase of internal sodium through opening of sodium channels, or with activation of other membrane sodium-transporting systems.
![Research paper thumbnail of Effects of Nitric Oxide Donors on Ca2+-Dependent [14C]GABA Release from Brain Synaptosomes: The Role of SH-Groups](https://attachments.academia-assets.com/42404138/thumbnails/1.jpg)
Biochemistry (Moscow)
Nitric oxide (NO) modulates processes of synaptic transmission at pre- and postsynaptic levels. I... more Nitric oxide (NO) modulates processes of synaptic transmission at pre- and postsynaptic levels. In the present work we studied the mechanisms of action of NO on [gamma-14C]amino-n-butyric acid ([14C]GABA) release in rat cortical synaptosomes. NO donors--S-nitroso-L-cysteine and hydroxylamine (but not sodium nitroprusside)--inhibited the neurotransmitter efflux in a concentration range from 10 microM to 1 mM. Nitrosocysteine completely and selectively suppressed the Ca2+-dependent (vesicular) [14C]GABA release, while not affecting the Ca2+-independent component of the [14C]GABA transport. The influence of NO donors was not related to activation of guanylyl cyclase, since the membrane-permeable cGMP analog dibutyryl-cGMP did not mimic and the guanylyl cyclase inhibitor methylene blue did not change the NO effects. In contrast, the membrane-permeable SH-reagent N-ethylmaleimide (NEM) resembled the effects of NO donors on the Ca2+-dependent [14C]GABA release. The degree of inhibition of...
Journal of Neurochemistry
Cellular and molecular biology (Noisy-le-Grand, France)
Decreased staining with dimethylthiazol diphenyltetrazolium (MTT) is widely used for cell death d... more Decreased staining with dimethylthiazol diphenyltetrazolium (MTT) is widely used for cell death detection. This study examined MTT assay as a marker of the Na+i,K+i-independent mode of cell death revealed in ouabain-treated C7-MDCK cells derived from distal tubule of the Madin-Darby canine kidney. The action of 3-M ouabain on MTT reduction in C7-MDCK cells exhibited bipartite kinetics with a rapid ~2-fold decline occurring in 30-120 min and a delayed ~8-10-fold decrease after 10 hr of ouabain addition. Treatment with ouabain for 18 hr led to 6-fold activation of caspase-3, 4-fold elevation of chromatin fragmentation, and massive cell detachment. Caspase-3 activation, chromatin fragmentation and cell detachment were completely abolished by acidification of the incubation medium from pH 7.2 to 6.7. In contrast, the 2-fold inhibition of MTT reduction seen in 5 hr of ouabain addition was not affected by medium acidification. Within the 5-hr time window, we did not observe any significan...
Volume-regulated anion channels (VRACs) are ubiquitously expressed in virtually all mammalian cel... more Volume-regulated anion channels (VRACs) are ubiquitously expressed in virtually all mammalian cells. VRACs are activated in response to cell swelling and are permeable towards Cl- and small organic osmolytes including the amino acids glutamate and taurine. Although the presumed major physiological role for VRACs is the maintenance of cell volume at a constant level, they are also thought to contribute to a variety of other processes suchas cell cycle progression, regulation of vascular tone and apoptosis. We speculate that in the brain astrocytic VRAC functions may include intercellular communication involving the release of excitatory amino acids (EAAs). A major obstacle to this idea is that, although moderate astrocytic swelling is seen under physiological conditions, it is not sufficient to substantially activate VRACs and VRAC-mediated EAA release. Here we report that a variety of neurotransmitters potently stimulate VRACs and EAA release. This will be illustrated by examples of...
Apoptosis, 2015
In rodents, ubiquitous α1-Na(+), K(+)-ATPase is inhibited by ouabain and other cardiotonic steroi... more In rodents, ubiquitous α1-Na(+), K(+)-ATPase is inhibited by ouabain and other cardiotonic steroids (CTS) at ~10(3)-fold higher concentrations than those effective in other mammals. To examine the specific roles of the CTS-sensitive α1S- and CTS-resistant α1R-Na(+), K(+)-ATPase isoforms, we compared the effects of ouabain on intracellular Na(+) and K(+) content, cell survival, and mitogen-activated protein kinases (MAPK) in human and rat vascular smooth muscle cells (HASMC and RASMC), human and rat endothelial cells (HUVEC and RAEC), and human and rat brain astrocytes. 6-h exposure of HASMC and HUVEC to 3 μM ouabain dramatically increased the intracellular [Na(+)]/[K(+)] ratio to the same extend as in RASMC and RAEC treated with 3000 μM ouabain. In 24, 3 μM ouabain triggered the death of all types of human cells used in this study. Unlike human cells, we did not detect any effect of 3000-5000 μM ouabain on the survival of rat cells, or smooth muscle cells from mouse aorta (MASMC). Unlike in the wild-type α1(R/R) mouse, ouabain triggered death of MASMC from α1(S/S) mouse expressing human α1-Na(+), K(+)-ATPase. Furthermore, transfection of HUVEC with rat α1R-Na(+), K(+)-ATPase protected them from the ouabain-induced death. In HUVEC, ouabain led to phosphorylation of p38 MAPK, whereas in RAEC it stimulated phosphorylation of ERK1/2. Overall, our results, demonstrate that the drastic differences in cytotoxic action of ouabain on human and rodent cells are caused by unique features of α1S/α1R-Na(+), K(+)-ATPase, rather than by any downstream CTS-sensitive/resistant components of the cell death machinery.
Biochemistry. Biokhimii͡a, 1998
Nitric oxide (NO) is known to potentiate neurotransmitter release in several types of neuronal ce... more Nitric oxide (NO) is known to potentiate neurotransmitter release in several types of neuronal cells. In the present study, the influence of NO on the membrane potential of isolated nerve endings (synaptosomes) from rat brain was studied. NO donors--sodium nitroprusside (SNP), S-nitroso-L-cysteine (CysNO), and hydroxylamine (HA)--induced synaptosome depolarization monitored by decreasing accumulation of 86Rb+ and the lipophilic potential-sensitive probe [3H]tetraphenylphosphonium. SNP reduced plasma membrane potential by 3-5 mV with half-maximal effect at approximately 10 microM. More potent NO donors, CysNO and HA, led to significant depolarization of the plasma membrane at 10-100 microM concentrations and also induced depolarization of mitochondria at concentrations above 1 mM. At 10 microM-10 mM concentrations, NO donors inhibited potassium channels; CysNO and HA also suppressed the activity of the sodium pump. NO-induced depolarization was not blocked by guanylate cyclase inhibi...
Acta neurobiologiae experimentalis, 1997
Neuromethods, 2014
Electrospun poly-L -lactic acid (PLLA) fi bers are presently explored as tissue engineering platf... more Electrospun poly-L -lactic acid (PLLA) fi bers are presently explored as tissue engineering platforms for regeneration of the central nervous system. In particular, aligned, electrospun fi bers are capable of directing astrocyte cellular extension and migration. The precise mechanisms by which aligned, electrospun substrates alter glial cell behavior are poorly understood. Therefore, there is a need for designing and refi ning electrospun fi ber platforms and developing novel approaches for studying astrocytic behavior and physiology on aligned substrates. Here, we describe and discuss methods for (1) fabrication of fl uorescent PLLA microfi bers by electrospinning, (2) coating PLLA fi bers with different extracellular matrix (ECM) proteins to facilitate attachment of astroglial cells, (3) isolation of primary astrocytes and plating them onto PLLA fi bers, and (4) imaging the interactions between PLLA fi bers and astrocytes to better understand the ability of fi bers to enable astrocyte extension and migration.
PLoS ONE, 2008
A variety of physiological and pathological factors induce cellular swelling in the brain. Change... more A variety of physiological and pathological factors induce cellular swelling in the brain. Changes in cell volume activate several types of ion channels, which mediate the release of inorganic and organic osmolytes and allow for compensatory cell volume decrease. Volume-regulated anion channels (VRAC) are thought to be responsible for the release of some of organic osmolytes, including the excitatory neurotransmitters glutamate and aspartate. In the present study, we compared the in vivo properties of the swelling-activated release of glutamate, aspartate, and another major brain osmolyte taurine. Cell swelling was induced by perfusion of hypoosmotic (low [NaCl]) medium via a microdialysis probe placed in the rat cortex. The hypoosmotic medium produced several-fold increases in the extracellular levels of glutamate, aspartate and taurine. However, the release of the excitatory amino acids differed from the release of taurine in several respects including: (i) kinetic properties, (ii) sensitivity to isoosmotic changes in [NaCl], and (iii) sensitivity to hydrogen peroxide, which is known to modulate VRAC. Consistent with the involvement of VRAC, hypoosmotic medium-induced release of the excitatory amino acids was inhibited by the anion channel blocker DNDS, but not by the glutamate transporter inhibitor TBOA or Cd 2+ , which inhibits exocytosis. In order to elucidate the mechanisms contributing to taurine release, we studied its release properties in cultured astrocytes and cortical synaptosomes. Similarities between the results obtained in vivo and in synaptosomes suggest that the swelling-activated release of taurine in vivo may be of neuronal origin. Taken together, our findings indicate that different transport mechanisms and/or distinct cellular sources mediate hypoosmotic mediuminduced release of the excitatory amino acids and taurine in vivo.
Free Radical Biology and Medicine, 2014
The contribution of oxidative stress to ischemic brain damage is well established. Nevertheless, ... more The contribution of oxidative stress to ischemic brain damage is well established. Nevertheless, for unknown reasons, several clinically tested antioxidant therapies have failed to show benefits in human stroke. Based on our previous in vitro work, we hypothesized that the neuroprotective potency of antioxidants is related to their ability to limit the release of the excitotoxic amino acids glutamate and aspartate. We explored the effects of two antioxidants, tempol and edaravone, on amino acid release in the brain cortex, in a rat model of transient occlusion of the middle cerebral artery (MCAo). Amino acid levels were quantified using a microdialysis approach, with the probe positioned in the ischemic penumbra as verified by a laser Doppler technique. Two-hour MCAo triggered a dramatic increase in the levels of glutamate, aspartate, taurine, and alanine. Microdialysate delivery of 10 mM tempol reduced the amino acid release by 60-80%, whereas matching levels of edaravone had no effect. In line with these data, an intracerebroventricular injection of tempol but not edaravone (500 nmol each, 15 min before MCAo) reduced infarction volumes by $50% and improved neurobehavioral outcomes. In vitro assays showed that tempol was superior at removing superoxide anion, whereas edaravone was more potent at scavenging hydrogen peroxide, hydroxyl radical, and peroxynitrite. Overall, our data suggest that the neuroprotective properties of tempol are probably related to its ability to reduce tissue levels of the superoxide anion and pathological glutamate release and, in such a way, limit progression of brain infarction within ischemic penumbra. These new findings may be instrumental in developing new antioxidant therapies for treatment of stroke.
The Journal of Physiology, 2006
Ubiquitously expressed volume-regulated anion channels (VRACs) are chloride channels which are pe... more Ubiquitously expressed volume-regulated anion channels (VRACs) are chloride channels which are permeable to a variety of small organic anions, including the excitatory amino acids (EAAs) glutamate and aspartate. Broad spectrum anion channel blockers strongly reduce EAA release in cerebral ischaemia and other pathological states associated with prominent astrocytic swelling. However, it is uncertain whether VRAC serves as a major pathway for EAA release from swollen cells. In the present study, we measured swelling-activated release of EAAs as D-[ 3 H]aspartate efflux, and VRAC-mediated Clcurrents by whole-cell patch clamp in cultured rat astrocytes. We compared the pharmacological profiles of the swelling-activated EAA release pathway and Clcurrents. The expression of candidate Clchannels was confirmed by RT-PCR. The maxi Clchannel (p-VDAC) blocker Gd 3+ , the ClC-2 inhibitor Cd 2+ , and the MDR-1 blocker verapamil did not affect EAA release or VRAC currents. An antagonist of calcium-sensitive Clchannels (CaCC), niflumic acid, had little effect on EAA release and only partially inhibited swelling-activated Clcurrents. The phorbol ester PDBu, which blocks ClC-3-mediated Clcurrents, had no effect on VRAC currents and up-regulated EAA release. In contrast, DCPIB, which selectively inhibits VRACs, potently suppressed both EAA release and VRAC currents. Two other relatively selective VRAC inhibitors, tamoxifen and phloretin, also blocked the VRAC currents and strongly reduced EAA release. Taken together, our data suggest that (i) astrocytic volume-dependent EAA release is largely mediated by the VRAC, and (ii) the ClC-2, ClC-3, ClC-4, ClC-5, VDAC, CaCC, MDR-1 and CFTR gene products do not contribute to EAA permeability.
Stroke, 2002
Stroke welcomes Letters to the Editor and will publish them, if suitable, as space permits. They ... more Stroke welcomes Letters to the Editor and will publish them, if suitable, as space permits. They should not exceed 750 words (excluding references) and may be subject to editing or abridgment. Please submit letters in duplicate, typed double-spaced. Include a fax number for the corresponding author and a completed copyright transfer agreement form (published in every other issue). 1. Vliegenthart R, Hollander M, Breteler MMB, van der Kuip DAM, Hofman A, Oudkerk M, Witteman JCM. Stroke is associated with coronary calcification as detected by electron-beam CT: the Rotterdam Coronary Calcification Study. Stroke. 2002;33:462-465. 2. Agatston AS, Janowitz WR, Hildner FJ, Zusmer NR, Viamonte M Jr, Detrano R. Quantification of coronary artery calcium using ultrafast computed tomography. J Am Coll Cardiol. 1990;15:827-832. 3. Becker CR, Jakobs TF, Aydemir S, Becker A, Knez A, Schoepf UJ, Bruening R, Haberl R, Reiser MF. Helical and single-slice conventional CT versus electron beam CT for the quantification of coronary artery calcification. AJR Am J Roentgenol. 2000;174:543-547. 4. Newman AB, Naydeck BL, Sutton-Tyrrell K, Feldman A, Edmundowicz D, Kuller LH. Coronary artery calcification in older adults to age 99: prevalence and risk factors.
PLoS ONE, 2010
Gliomas are morbid brain tumors that are extremely resistant to available chemotherapy and radiol... more Gliomas are morbid brain tumors that are extremely resistant to available chemotherapy and radiology treatments. Some studies have suggested that calcium-activated potassium channels contribute to the high proliferative potential of tumor cells, including gliomas. However, other publications demonstrated no role for these channels or even assigned them antitumorogenic properties. In this work we characterized the expression and functional contribution to proliferation of Ca 2+ -activated K + channels in human glioblastoma cells. Quantitative RT-PCR detected transcripts for the big conductance (BK), intermediate conductance (IK1), and small conductance (SK2) K + channels in two glioblastoma-derived cell lines and a surgical sample of glioblastoma multiforme. Functional expression of BK and IK1 in U251 and U87 glioma cell lines and primary glioma cultures was verified using whole-cell electrophysiological recordings. Inhibitors of BK (paxilline and penitrem A) and IK1 channels (clotrimazole and TRAM-34) reduced U251 and U87 proliferation in an additive fashion, while the selective blocker of SK channels UCL1848 had no effect. However, the antiproliferative properties of BK and IK1 inhibitors were seen at concentrations that were higher than those necessary to inhibit channel activity. To verify specificity of pharmacological agents, we downregulated BK and IK1 channels in U251 cells using gene-specific siRNAs. Although siRNA knockdowns caused strong reductions in the BK and IK1 current densities, neither single nor double gene silencing significantly affected rates of proliferation. Taken together, these results suggest that Ca 2+ -activated K + channels do not play a critical role in proliferation of glioma cells and that the effects of pharmacological inhibitors occur through their off-target actions.
PLoS ONE, 2013
Cutaneous ATP release plays an important role in both epidermal stratification and chronic pain, ... more Cutaneous ATP release plays an important role in both epidermal stratification and chronic pain, but little is known about ATP release mechanisms in keratinocytes that comprise the epidermis. In this study, we analyzed ATP release from cultured human neonatal keratinocytes briefly exposed to air, a process previously demonstrated to trigger ATP release from these cells. We show that exposing keratinocytes to air by removing media for 15 seconds causes a robust, long-lasting ATP release. This air-stimulated ATP release was increased in calcium differentiated cultures which showed a corresponding increase in connexin 43 mRNA, a major component of keratinocyte hemichannels. The known connexin hemichannel inhibitors 1-octanol and carbenoxolone both significantly reduced air-stimulated ATP release, as did two drugs traditionally used as ABC transporter inhibitors (glibenclamide and verapamil). These same 4 inhibitors also prevented an increase in the uptake of a connexin permeable dye induced by air exposure, confirming that connexin hemichannels are open during airstimulated ATP release. In contrast, activity of the MDR1 ABC transporter was reduced by air exposure and the drugs that inhibited air-stimulated ATP release had differential effects on this transporter. These results indicate that air exposure elicits non-vesicular release of ATP from keratinocytes through connexin hemichannels and that drugs used to target connexin hemichannels and ABC transporters may cross-inhibit. Connexins represent a novel, peripheral target for the treatment of chronic pain and dermatological disease. Citation: Barr TP, Albrecht PJ, Hou Q, Mongin AA, Strichartz GR, et al. (2013) Air-Stimulated ATP Release from Keratinocytes Occurs through Connexin Hemichannels. PLoS ONE 8(2): e56744.
Pathophysiology, 2001
In animal organisms, cell volume undergoes dynamic changes in many physiological and pathological... more In animal organisms, cell volume undergoes dynamic changes in many physiological and pathological processes. To protect themselves against lysis and apoptosis and to maintain an optimal concentration of intracellular enzymes and metabolites, most animal cells actively regulate their volume. In the present review, we shortly summarize the data on ion transport mechanisms involved in regulatory volume decrease (RVD) and regulatory volume increase (RVI) with an emphasis on unresolved aspects of this problem such as: (i) how cells sense their volume changes; (ii) what signals are generated upon cell volume alterations; and (iii) how these signals are transferred to the ion transport systems executing cell volume regulation.
Neurochemistry International, 1997
Influence of hypotonic swelling on Ca 2 ÷ (45Ca2+) uptake in rat brain synaptosomes was studied. ... more Influence of hypotonic swelling on Ca 2 ÷ (45Ca2+) uptake in rat brain synaptosomes was studied. A decrease in medium osmolality from 310 to 260-180 mOsm led to a progressive stimulation of 45Ca2 ÷ accumulation. The effect was blocked by verapamil (IC50 = 5/~M), COC12 (ICs0 = 58/~M) and retained at a fixed concentration of external sodium indicating the involvement of Ca 2 ÷ channels rather than Na ÷/Ca 2+ exchange in swelling-induced Ca 2+ influx. The populations of calcium channels observed in hypoosmotic and depolarizing conditions are different in three aspects: (i) kinetics of 45Ca2+ entry; (ii) insensitivity to dihydropyridines and co-conotoxin GVIA; (iii) insensitivity to preliminary depolarization by high potassium. The effects of swelling and depolarization on Ca 2+ uptake were additive. No change in membrane potential monitored with diS-C3-(5) was recorded during synaptosome hypotonic swelling. The results suggest the existence in synaptosomal plasma membrane of volume-dependent calcium-permeable channels with properties distinct from those of the voltage-dependent calcium channels. Activation of these channels may constitute an early event in volume regulation of nerve terminals in anisoosmotic conditions.
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Papers by Alexander Mongin