Activation of Cl-dependent K transport in Ehrlich ascites tumor cells

1986 ◽  
Vol 251 (3) ◽  
pp. C369-C379 ◽  
Author(s):  
B. Kramhoft ◽  
I. H. Lambert ◽  
E. K. Hoffmann ◽  
F. Jorgensen
Keyword(s):  
Regulatory Volume Decrease ◽  
Extracellular Ph ◽  
Cell Swelling ◽  
Disulfonic Acid ◽  
Cell Shrinkage ◽  
Ehrlich Cells ◽  
Ehrlich Ascites ◽  
Dependent Component ◽  
Ca Depletion ◽  
Volume Decrease

N-ethylmaleimide (NEM) treatment of steady-state Ehrlich cells induces a substantial net loss of cellular KCl and cell shrinkage. The majority of the initial K loss is Cl dependent. From estimates of membrane potential it is concluded that the NEM-induced KCl loss is electroneutral. The effect of NEM on H extrusion by cells in 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS)-containing medium showed that only an insignificant part of the K loss could be attributed to an activation of a K-H exchange system. Consequently, NEM appears to activate a K-Cl cotransport, which causes cell shrinkage. The anion preference of the K loss is Cl greater than Br much greater than SCN = NO3. NEM also seems to inhibit a Cl-dependent Na uptake previously described in shrunken cells. Addition of NEM to cells undergoing regulatory volume decrease after swelling in hyposmotic media results in a Cl-dependent acceleration of cell shrinkage, suggesting that a Cl-dependent component of K efflux is induced by NEM also in swollen cells. A Cl-dependent K efflux is also activated in Ca-depleted cells or at reduced extracellular pH after cell swelling. Under isotonic conditions activation of Cl-dependent K flux after Ca depletion or pH reduction could not be demonstrated. The combined results show that Ehrlich cells possess a latent K-Cl cotransport that becomes active after changes in the state of SH groups, regardless of the initial cell volume. A similar K-Cl cotransport is activated in hypotonically swollen cells after Ca depletion or after reduction of the extracellular pH.

scholarly journals Vasopressin Neurons Respond to Hyperosmotic Stimulation with Regulatory Volume Increase and Secretory Volume Decrease by Activating Ion Transporters and Ca2+ Channels

2021 ◽  
Vol 55 (S1) ◽  
pp. 119-134
Keyword(s):  
Regulatory Volume Decrease ◽  
The Body ◽  
Cell Swelling ◽  
Ion Transporters ◽  
Cell Shrinkage ◽  
Volume Increase ◽  
Regulatory Volume Increase ◽  
Vasopressin Neurons ◽  
Ca2 Channels ◽  
Volume Decrease

BACKGROUND/AIMS: Arginine vasopressin (AVP) neurons play an important role for sensing a change in the plasma osmolarity and thereby responding with regulated AVP secretion in order to maintain the body fluid homeostasis. The osmo-sensing processes in magnocellular neurosecretory cells (MNCs) including AVP and oxytocin (OXT) neurons of the hypothalamus were reported to be coupled to sustained osmotic shrinkage or swelling without exhibiting discernible cell volume regulation. Since increasing evidence has shown some important differences in properties between AVP and OXT neurons, osmotic volume responses are to be reexamined with distinguishing these cell types from each other. We previously reported that AVP neurons identified by transgenic expression of enhanced green fluorescence protein (eGFP) possess the ability of regulatory volume decrease (RVD) after hypoosmotic cell swelling. Thus, in the present study, we examined the ability of regulatory volume increase (RVI) after hyperosmotic cell shrinkage in AVP neurons. METHODS: Here, we used eGFP-identified AVP neurons acutely dissociated from AVP-eGFP transgenic rats. We performed single-cell size measurements, cytosolic RT-PCR analysis, AVP secretion measurements, and patch-clamp studies. RESULTS: The AVP neurons were found to respond to a hyperosmotic challenge with physiological cell shrinkage caused by massive secretion of AVP, called a secretory volume decrease (SVD), superimposed onto physical osmotic cell shrinkage, and also to exhibit the ability of RVI coping with osmotic and secretory cell shrinkage. Furthermore, our pharmacological and molecular examinations indicated that AVP secretion and its associated SVD event are triggered by activation of T-type Ca2+ channels, and the RVI event is attained by parallel operation of Na+/H+ exchanger and Cl-/HCO3- anion exchanger. CONCLUSION: Thus, it is concluded that AVP neurons respond to hyperosmotic stimulation with the regulatory volume increase and the secretory volume increase by activating ion transporters and Ca2+ channels, respectively.

Regulatory volume decrease in lamprey erythrocytes: mechanisms of K+ and Cl- loss

1995 ◽  
Vol 268 (3) ◽  
pp. R590-R597
Author(s):  
L. V. Virkki ◽  
M. Nikinmaa
Keyword(s):  
Acetic Acid ◽  
Regulatory Volume Decrease ◽  
Cell Swelling ◽  
Disulfonic Acid ◽  
Minor Effect ◽  
Transport Pathways ◽  
Cl Transport ◽  
Medium Osmolarity ◽  
A Minor ◽  
Volume Decrease

The nature of the swelling-activated K+ and Cl- transport pathways of lamprey (Lampetra fluviatilis) erythrocytes was studied. In isosmotic medium, unidirectional K+ and Cl- effluxes appear to be largely mediated by conductive pathways. Unidirectional Cl- efflux increased as a function of a decrease in medium osmolarity. The swelling-activated Cl- transport was inhibited by R(+)-[(2-n-butyl-6,7-dichloro-2-cyclopentyl-2,3-dihydro-1-oxo-1H-inde n-5- yl)oxy]acetic acid (DIOA), furosemide, and 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). In contrast, moderate cell swelling did not increase unidirectional ouabain-insensitive K+ efflux. However, inhibition of transport by Ba2+ was markedly reduced. This suggests that the Ba(2+)-sensitive pathway that mediated most of the K+ efflux in isosmotic conditions was inhibited by cell swelling and a Ba(2+)-insensitive pathway was activated. DIOA had no effect on K+ efflux in isosmotic or hyposmotic medium. These data and the finding that substitution of NO3- or SCN- for Cl- had only a minor effect on the swelling-induced net extrusion of K+ and water indicate that the pathways for K+ and Cl-, activated by cell swelling, are conductive.

Separate K+ and Cl- transport pathways are activated for regulatory volume decrease in jejunal villus cells

1991 ◽  
Vol 260 (3) ◽  
pp. G405-G415 ◽  
Author(s):  
R. J. MacLeod ◽  
J. R. Hamilton
Keyword(s):  
Regulatory Volume Decrease ◽  
Cell Swelling ◽  
Disulfonic Acid ◽  
Transport Pathways ◽  
Kg Medium ◽  
Cell Sizing ◽  
Rate Limiting ◽  
Li Substitution ◽  
Hypotonic Swelling ◽  
Volume Decrease

We assessed ion transport mechanisms operative during regulatory volume decrease (RVD) in jejunal villus enterocytes, isolated in suspension from guinea pig jejunum and examined with electronic cell sizing. Immediately after reduction of osmolarity (153 mosmol/kg medium) enterocytes swelled, but within 5 min they shrank by 50%. This RVD, which was complete by 20 min, was unaffected by Li+ substitution for Na+ or by Na(+)-free (N-methyl-D-glucose, NMDG+) medium. Passive loss of K+ is required for RVD because both the magnitude and direction of RVD changed when external [K+] varied. Increasing K+ permeability with gramicidin (0.5 microM) accelerated RVD in NMDG+ medium (10.0 +/- 0.8 vs. 6.2 +/- 0.4% min-1, P less than 0.01) suggesting that K+ loss is rate limiting for RVD. Inhibition of K(+)- and Ca2(+)-activated K+ conductance with Ba2+ (5 mM, P less than 0.005), quinine (100 microM, P less than 0.005), or apamin (1 microM, P less than 0.005) prevented RVD. Inhibition of Cl- conductance with 9-anthracenecarboxylic acid (100 microM, P less than 0.005) or dipyridamole (75 microM, P less than 0.005) also prevented RVD. In isotonic HCO3(-)-buffered medium, the addition of gramicidin to cells generated conditions in which anion permeability was rate limiting for cell swelling. This swelling was inhibited 97% by 100 microM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). In K(+)-free, HCO3(-)-buffered medium containing DIDS and gramicidin hypotonic swelling resulted in continued (secondary) swelling (rel vol 1.19 +/- 0.01 vs. 1.25 +/- 0.02, P less than 0.02).(ABSTRACT TRUNCATED AT 250 WORDS)

Volume-activated K+ and Cl- pathways of dissociated epithelial cells (MDCK): role of Ca2+

1990 ◽  
Vol 258 (5) ◽  
pp. C827-C834 ◽  
Author(s):  
A. Rothstein ◽  
E. Mack
Keyword(s):  
Mdck Cells ◽  
Regulatory Volume Decrease ◽  
Disulfonic Acid ◽  
Ionophore A23187 ◽  
Madin Darby Canine Kidney ◽  
Osmotic Swelling ◽  
Volume Activation ◽  
Darby Canine Kidney ◽  
Volume Decrease

Osmotic swelling of dissociated Madin-Darby canine kidney (MDCK) cells in NaCl medium is followed by shrinking (regulatory volume decrease, or RVD) or in KCl medium by secondary swelling. The cation ionophore gramicidin has little effect on volumes of isotonic cells but accelerates volume-activated changes in either medium. Immediately after hypotonic exposure, the membrane becomes transiently hyperpolarized followed by depolarization. The depolarization phase is diminished by the anion transport inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). Swelling is also associated with an almost immediate increase in Ca2+ influx and elevation of cytoplasmic Ca2+ ([Ca2+]i) preceding RVD. In Ca2(+)-free medium, [Ca2+]i rapidly declines to a low level. Osmotic swelling, under these circumstances, is associated with a small transient increase in [Ca2+]i, but RVD or secondary swelling (in KCl) are minimal. Under these conditions, addition of gramicidin or the Ca2(+)-ionophore A23187 induces significant volume changes, although not as large as those found in the presence of Ca2+. Quinine inhibits RVD in the absence of gramicidin, but not in its presence; oligomycin C, DIDS, and trifluoperazine, on the other hand, inhibit in the presence of the ionophore. These findings suggest that in MDCK cells RVD involves activation of distinct conductive K+ and Cl- pathways which allow escape of KCl and osmotically obligated water and that activation of both pathways is associated with elevated [Ca2+]i derived largely from volume activation of a Ca2(+)-influx pathway.

Resistance to osmotic lysis in BXD-31 mouse erythrocytes: association with upregulated K-Cl cotransport

1996 ◽  
Vol 270 (3) ◽  
pp. C866-C877 ◽  
Author(s):  
C. C. Armsby ◽  
A. K. Stuart-Tilley ◽  
S. L. Alper ◽  
C. Brugnara
Keyword(s):  
Normal Cell ◽  
Genetic Locus ◽  
Regulatory Volume Decrease ◽  
Osmotic Fragility ◽  
Cell Swelling ◽  
K Channel ◽  
Cell Dehydration ◽  
Osmotic Lysis ◽  
Volume Decrease ◽  
Characteristic Resistance

The decreased osmotic fragility and reduced K+ content of BXD-31 mouse erythrocytes arise from variation at a single genetic locus. We compared ion transport in erythrocytes from BXD-31 mice and the parental strain, DBA/2J. The strains had similar rates for Na-K pump, Na/H exchange, Na-K-2Cl cotransport, Ca2+ activated K+ channel, or AE1-mediated SO4 transport. In contrast, K-Cl cotransport was twice as active in BXD-31 as in DBA/2J cells. Cl- dependent K+ efflux from BXD-31 cells displayed steep activation by acid pH (with maximal transport occurring at pH 6.75), whereas DBA/2J erythrocytes displayed a far less dramatic response to pH. Both strains displayed regulatory volume decrease in response to cell swelling. However, a 62% greater loss of cell K+ via K-Cl cotransport was observed in the BXD-31 strain. Furthermore the decreased osmotic fragility of BXD-31 red blood cells was normalized by treatment with nystatin to achieve normal cell K+ and water content. Thus upregulated K-Cl cotransport induces cell dehydration and K+ deficit in BXD-31 erythrocytes and causes their characteristic resistance to osmotic lysis.

Regulatory volume decrease in HL-60 cells: importance of rapid changes in permeability of Cl- and organic solutes

1994 ◽  
Vol 267 (4) ◽  
pp. C1045-C1056 ◽  
Author(s):  
K. R. Hallows ◽  
P. A. Knauf
Keyword(s):  
Regulatory Volume Decrease ◽  
Organic Osmolytes ◽  
Diffusion Kinetic ◽  
Nernst Potential ◽  
Ehrlich Ascites ◽  
Rapid Changes ◽  
36Cl Fluxes ◽  
Free Media ◽  
Rate Limiting ◽  
Volume Decrease

Results obtained through the use of inhibitors and isotope flux and equilibration techniques indicate that the regulatory volume decrease (RVD) response of human promyelocytic leukemic HL-60 cells occurs largely through the efflux of K+ and Cl- through separate conductive membrane pathways. These "channels" differ pharmacologically and in their modes of activation from those described in lymphocytes and Ehrlich ascites tumor cells. With use of measured 86Rb+ and 36Cl- fluxes, together with a diffusion kinetic model, the membrane potential (Em) and apparent K+ and Cl- permeabilities (PK and PCl) were estimated under various isotonic and hypotonic conditions. Under isotonic (300 mosM) conditions, Em is close to the Nernst potential for K+ and PCl is < 0.1 PK. Rapid and steeply graded increases in the measured Cl- efflux rate and calculated PCl occur with decreasing tonicity, with the largest increases at tonicities < 80% of isotonic. K+ efflux and the apparent PK increase only modestly with decreasing tonicity. At 50% tonicity, PCl rises to nearly 10 times PK, which should cause substantial membrane depolarization, with Em approaching the Nernst potential for Cl-. Gramicidin treatment markedly accelerates the rate of RVD and net 36Cl- efflux in hypotonic Na(+)-and Cl(-)-free media, providing further evidence that PK is rate limiting during RVD. K+ loss exceeds Cl- loss during RVD, and the total loss of K+ and Cl- is insufficient to account for the observed degree of volume recovery in 50% tonicity media, indicating that other (organic) osmolytes must take part in the HL-60 cell RVD response.

Control of intracellular pH during regulatory volume decrease in HL-60 cells

1994 ◽  
Vol 267 (4) ◽  
pp. C1057-C1066 ◽  
Author(s):  
K. R. Hallows ◽  
D. Restrepo ◽  
P. A. Knauf
Keyword(s):  
Intracellular Ph ◽  
Ph Regulation ◽  
Regulatory Volume Decrease ◽  
Transport Systems ◽  
Disulfonic Acid ◽  
Volume Dependence ◽  
Supportive Role ◽  
22Na Uptake ◽  
Volume Decrease ◽  
Stimulation Of

Intracellular pH (pHi) homeostasis was investigated in human promyelocytic leukemic HL-60 cells as they undergo regulatory volume decrease (RVD) in hypotonic media to determine how well pHi is regulated and which transport systems are involved. Cells suspended in hypotonic (50-60% of isotonic) media undergo a small (< 0.2 pH units), but significant (P < 0.05), intracellular acidification within 5 min. However, after 30 min of RVD, pHi is not significantly different from the initial pHi in 20 mM HCO3- medium and is significantly higher in HCO3(-)-free medium. Experiments performed in media with or without 150 microM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid and HCO3- demonstrate that the anion exchanger (AE) mediates a net Cl- influx, with compensating HCO3- efflux, during RVD. To determine which transport systems are involved in counteracting this tendency toward acidification, we measured transport rates and examined the effect of transport system inhibitors on pHi. We found that inhibition of Na+/H+ exchange (NHE) with 12.5 microM ethylisoproplamiloride (EIPA) causes pHi to fall significantly by the end of 30 min of RVD. As assessed by EIPA-sensitive 22Na+ uptake measurements, NHE, largely dormant under resting isotonic conditions, becomes significantly activated by the end of 30 min of RVD, despite recovery of pHi and cell volume to near-normal levels. Thus a shift in the normal pHi dependence and/or volume dependence of NHE activity must occur during RVD under hypotonic conditions. In contrast, H(+)-monocarboxylate cotransport appears to play only a supportive role in pH regulation during RVD, as indicated by lack of stimulation of [14C]lactate efflux during RVD.

Physiological significance of hypotonicity-induced regulatory volume decrease: reduction in intracellular Cl− concentration acting as an intracellular signaling

2007 ◽  
Vol 292 (5) ◽  
pp. F1411-F1417 ◽  
Author(s):  
Hiroaki Miyazaki ◽  
Atsushi Shiozaki ◽  
Naomi Niisato ◽  
Yoshinori Marunaka
Keyword(s):  
Intracellular Signaling ◽  
Channel Blocker ◽  
Fluorescent Dyes ◽  
Regulatory Volume Decrease ◽  
Cell Swelling ◽  
Dependent Manner ◽  
Stimulatory Action ◽  
A6 Cells ◽  
Intracellular Signal ◽  
Volume Decrease

Regulatory volume decrease (RVD) occurs after hypotonicity-caused cell swelling. RVD is caused by activation of ion channels and transporters, which cause effluxes of K+, Cl−, and H2O, leading to cell shrinkage. Recently, we showed that hypotonicity stimulated transepithelial Na+ reabsorption via elevation of epithelial Na+ channel (α-ENaC) expression in renal epithelia A6 cells in an RVD-dependent manner and that reduction of intracellular Cl− concentration ([Cl−]i) stimulated the Na+ reabsorption. These suggest that RVD would reveal its stimulatory action on the Na+ reabsorption by reducing [Cl−]i. However, the reduction of [Cl−]i during RVD has not been definitely analyzed due to technical difficulties involved in halide-sensitive fluorescent dyes. In the present study, we developed a new method for the measurement of [Cl−]i change during RVD by using a high-resolution flow cytometer with a halide-specific fluorescent dye, N-(6-methoxyquinolyl) acetoethyl ester. The [Cl−]i in A6 cells in an isotonic medium was 43.6 ± 3.1 mM. After hypotonic shock (268 to 134 mosmol/kgH2O), a rapid increase of cell volume followed by RVD occurred. The RVD caused drastic diminution of [Cl−]i from 43.6 to 10.8 mM. Under an RVD-blocked condition with NPPB (Cl− channel blocker) or quinine (K+ channel blocker), we did not detect the reduction of [Cl−]i. Based on these observations, we conclude that one of the physiological significances of RVD is the reduction of [Cl−]i and that RVD shows its action via reduction of [Cl−]i acting as an intracellular signal regulating cellular physiological functions.

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