Volume-sensitive anion channels mediate swelling-activated inositol and taurine efflux

1993 ◽  
Vol 265 (6) ◽  
pp. C1489-C1500 ◽  
Author(s):  
P. S. Jackson ◽  
K. Strange
Keyword(s):  
Fatty Acids ◽  
Anion Channel ◽  
Anion Transport ◽  
Cell Swelling ◽  
C6 Glioma Cells ◽  
Organic Osmolytes ◽  
Transport Pathway ◽  
Whole Cell ◽  
Efflux Mechanism ◽  
Taurine Efflux

C6 glioma cells accumulate the organic osmolyte inositol in response to chronic hypertonic stress. Upon return to isotonic conditions, cell swelling activates a Na(+)-independent passive low-affinity inositol efflux mechanism that is inhibited 80-100% by a number of anion transport blockers, certain lipoxygenase blockers, and various polyunsaturated fatty acids. Taurine efflux is also enhanced by cell swelling. The taurine efflux pathway has characteristics that are identical to those of the inositol efflux mechanism, including kinetics of activation and inactivation, osmotic sensitivity, pharmacological sensitivity, and inhibition by certain Na+ and Cl- substitutes. These results suggest strongly that volume-sensitive inositol and taurine efflux are mediated by a common transport mechanism. The inhibition of the transport pathway by anion transport blockers and unsaturated fatty acids suggests indirectly that efflux of these solutes may be mediated by an anion channel. Whole cell patch clamp measurements in CsCl solutions were used to test this hypothesis. Under hypertonic conditions, C6 cells had an extremely low membrane conductance (approximately 0.02 nS/pF). After cell swelling, however, whole cell anion conductance was activated rapidly to values up to 1.5-2 nS/pF. This conductance was outwardly rectified and selective for anions and was inhibited 80-100% by blockers of swelling-activated inositol and taurine efflux. The relative taurine permeability (i.e., Ptaurine/PCl) of the conductance was 0.20. Isosmotic replacement of raffinose in the external medium with inositol or sorbitol induced a transient inward current, suggesting that Cl- and these polyols compete for common binding sites on the channel. We conclude that a volume-sensitive anion channel mediates the efflux of structurally diverse organic osmolytes such as taurine and inositol from the cell.

The volume-sensitive organic osmolyte-anion channel VSOAC is regulated by nonhydrolytic ATP binding

1994 ◽  
Vol 267 (5) ◽  
pp. C1203-C1209 ◽  
Author(s):  
P. S. Jackson ◽  
R. Morrison ◽  
K. Strange
Keyword(s):  
Ketone Bodies ◽  
Anion Channel ◽  
Metabolic Inhibitors ◽  
Cell Swelling ◽  
Organic Osmolytes ◽  
Pipette Solution ◽  
Organic Osmolyte ◽  
Atp Levels ◽  
Patch Pipette ◽  
Taurine Efflux

Efflux of intracellular organic osmolytes to the external medium is a ubiquitous response to cell swelling. Accumulating evidence indicates that volume regulatory loss of structurally unrelated organic osmolytes from cells is mediated by a relatively nonselective volume-sensitive anion channel. In C6 cells, we have termed this channel VSOAC for volume-sensitive organic osmolyte-anion channel. Swelling-induced activation of VSOAC required the presence of ATP or nonhydrolyzable ATP analogues [adenosine 5'-O-(3-thiotriphosphate), adenylylmethyl-enediphosphonate (AMP-PCP), or 5'-adenylylimidodiphosphate] in the patch pipette. Sustained activation of VSOAC also required ATP. Channel rundown was observed when cellular ATP levels were lowered by intracellular dialysis with the patch pipette solution. Rundown was prevented by the ATP analogue AMP-PCP. Passive swelling-induced myo-[3H]inositol and [3H]taurine efflux was blocked by metabolic inhibitors that decreased cellular ATP levels. Titration of cellular ATP levels with azide demonstrated that the apparent dissociation constant (Kd) for ATP of both myo-inositol and taurine efflux was approximately 1.7 mM. The high Kd for ATP indicates that cellular metabolic state plays an important role in modulating organic osmolyte loss. Regulation of VSOAC activity by ATP prevents depletion of metabolically expensive organic osmolytes when cellular energy production is reduced. In addition, ATP-dependent regulation provides essential feedback to minimize the loss of energy-producing carbon sources such as pyruvate, short-chain fatty acids, ketone bodies, and amino acids, which readily permeate this channel.

Intracellular electrolytes regulate the volume set point of the organic osmolyte/anion channel VSOAC

1997 ◽  
Vol 272 (6) ◽  
pp. C1766-C1775 ◽  
Author(s):  
F. Emma ◽  
M. McManus ◽  
K. Strange
Keyword(s):  
Anion Channel ◽  
Cell Swelling ◽  
C6 Glioma Cells ◽  
Organic Osmolytes ◽  
Intact Cells ◽  
Hypertonic Medium ◽  
Set Point ◽  
Intracellular Electrolytes ◽  
Organic Osmolyte ◽  
Volume Sensitivity

Regulation of the volume sensitivity of the swelling-activated organic osmolyte/anion channel VSOAC by intracellular electrolytes was examined in intact and patch-clamped C6 glioma cells. In intact cells, VSOAC activation was monitored by [3H]taurine efflux measurements, and intracellular electrolyte concentrations were manipulated by acclimation to hypertonic medium for varying periods of time. Hypertonic shrinkage was followed by a rapid and complete regulatory volume increase mediated by electrolyte accumulation that elevated intracellular Na+, K+, and Cl- concentrations. During prolonged (4-48 h) exposure to hypertonicity, electrolyte concentrations decreased gradually as cells accumulated the organic osmolyte myo-inositol. VSOAC activation induced by cell swelling of 35-40% increased as a function of the time of exposure to hypertonicity and was inversely correlated with measured intracellular Na+, K+ and Cl- levels. In patch-clamped cells, swelling-induced Cl- current activation was unaffected by acclimation conditions but was inhibited by increasing the concentration of electrolytes in the patch pipette solution. Quantification of the relationship between VSOAC activation and cell swelling demonstrated that increases in intracellular electrolyte levels increase VSOAC volume set point. Regulation of VSOAC volume sensitivity by electrolytes allows cells to selectively utilize electrolytes or a combination of electrolytes and organic osmolytes for regulatory volume decrease (RVD). Control over the type of solute used for volume regulation is advantageous, allowing cells to control intracellular ionic composition and prevent increases in cytoplasmic ionic strength during RVD.

More than just a pressure relief valve: physiological roles of volume-regulated LRRC8 anion channels

2019 ◽  
Vol 400 (11) ◽  
pp. 1481-1496 ◽  
Author(s):  
Lingye Chen ◽  
Benjamin König ◽  
Tianbao Liu ◽  
Sumaira Pervaiz ◽  
Yasmin S. Razzaque ◽  
...  
Keyword(s):  
Volume Regulation ◽  
Regulatory Volume Decrease ◽  
Anion Channel ◽  
Therapy Resistance ◽  
Cell Swelling ◽  
Organic Osmolytes ◽  
Anion Channels ◽  
Relief Valve ◽  
Wide Range ◽  
And Migration

Abstract The volume-regulated anion channel (VRAC) is a key player in the volume regulation of vertebrate cells. This ubiquitously expressed channel opens upon osmotic cell swelling and potentially other cues and releases chloride and organic osmolytes, which contributes to regulatory volume decrease (RVD). A plethora of studies have proposed a wide range of physiological roles for VRAC beyond volume regulation including cell proliferation, differentiation and migration, apoptosis, intercellular communication by direct release of signaling molecules and by supporting the exocytosis of insulin. VRAC was additionally implicated in pathological states such as cancer therapy resistance and excitotoxicity under ischemic conditions. Following extensive investigations, 5 years ago leucine-rich repeat-containing family 8 (LRRC8) heteromers containing LRRC8A were identified as the pore-forming components of VRAC. Since then, molecular biological approaches have allowed further insight into the biophysical properties and structure of VRAC. Heterologous expression, siRNA-mediated downregulation and genome editing in cells, as well as the use of animal models have enabled the assessment of the proposed physiological roles, together with the identification of new functions including spermatogenesis and the uptake of antibiotics and platinum-based cancer drugs. This review discusses the recent molecular biological insights into the physiology of VRAC in relation to its previously proposed roles.

scholarly journals Volume-dependent taurine release from cultured astrocytes requires permissive [Ca2+]iand calmodulin

1999 ◽  
Vol 277 (4) ◽  
pp. C823-C832 ◽  
Author(s):  
Alexander A. Mongin ◽  
Zhaohui Cai ◽  
Harold K. Kimelberg
Keyword(s):  
Cell Swelling ◽  
Transport Systems ◽  
Channel Blockers ◽  
Organic Osmolytes ◽  
Taurine Release ◽  
Cultured Astrocytes ◽  
High K ◽  
Intracellular Ca ◽  
Free Media ◽  
Taurine Efflux

Cell swelling results in regulatory activation of multiple conductive anion pathways permeable toward a broad spectrum of intracellular organic osmolytes. Here, we explore the involvement of extracellular and intracellular Ca2+ in volume-dependent [3H]taurine efflux from primary cultured astrocytes and compare the Ca2+ sensitivity of this efflux in slow (high K+ medium induced) and fast (hyposmotic medium induced) cell swelling. Neither Ca2+-free medium nor Ca2+-channel blockers prevented the volume-dependent [3H]taurine release. In contrast, loading cells with the membrane-permeable Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane- N, N, N′, N′-tetraacetic acid (BAPTA)-AM suppressed [3H]taurine efflux by 65–70% and 25–30% under high-K+ and hyposmotic conditions, respectively. Fura 2 measurements confirmed that BAPTA-AM, but not Ca2+-free media, significantly reduced resting intracellular Ca2+concentration ([Ca2+]i). The calmodulin antagonists trifluoperazine and fluphenazine reversibly and irreversibly, respectively, inhibited the high-K+-induced [3H]taurine release, consistent with their known actions on calmodulin. In hyposmotic conditions, the effects were less pronounced. These data suggest that volume-dependent taurine release requires minimal basal [Ca2+]iand involves calmodulin-dependent step(s). Quantitative differences in Ca2+/calmodulin sensitivity of high-K+-induced and hyposmotic medium-induced taurine efflux are due to both the effects of the inhibitors on high-K+-induced cell swelling and their effects on transport systems and/or signaling mechanisms determining taurine efflux.

Taurine transport in skate hepatocytes. II. Volume activation, energy, and sulfhydryl dependence

1992 ◽  
Vol 262 (3) ◽  
pp. G451-G460 ◽  
Author(s):  
N. Ballatori ◽  
J. L. Boyer
Keyword(s):  
Plasma Membrane ◽  
Concentration Gradient ◽  
Regulatory Volume Decrease ◽  
Metabolic Inhibitors ◽  
Cell Swelling ◽  
Disulfonic Acid ◽  
Organic Osmolytes ◽  
Taurine Transport ◽  
Taurine Concentration ◽  
Taurine Efflux

Isolated skate (Raja erinacea) hepatocytes swollen in hypotonic media exhibited a regulatory volume decrease (RVD) that was associated with only a small increase in K+ or 86Rb+ efflux but a substantial increase in the release of taurine, an amino acid found in high concentrations in skate hepatocytes. Taurine efflux was stimulated in media made hypotonic by addition of H2O or removal of NaCl, as well as in cells swollen in isotonic media containing rapidly penetrating solutes (202 mM ethylene glycol or 202 mM additional urea substituted for 101 mM NaCl), suggesting that cell swelling rather than hyposmolarity is the stimulus for the activation of taurine release. In contrast, release of glutathione, L-[14C]alanine and other alpha-amino acids (e.g., threonine, serine, glutamate, glutamine, glycine, or valine) was unaffected by dilution with 40% H2O. Taurine efflux was not altered by replacement of extracellular Na+ with choline+ or K+ and was only slightly diminished by replacing Cl- with NO3-. Addition of 50 mM taurine or hypotaurine to the incubation media also had no effect on volume-stimulated [14C]taurine efflux, suggesting that the taurine concentration gradient across the plasma membrane is not the driving force. Volume-stimulated taurine transport was temperature sensitive, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid inhibitable (0.5 mM), and nearly completely blocked by metabolic inhibitors (2,4-dinitrophenol, KCN, sodium azide, oligomycin, carbonyl cyanide m-chlorophenylhydrazone, and antimycin A), suggesting an active energy-dependent process. Sulfhydryl-reactive reagents (N-ethylmaleimide, diamide, iodoacetate, tert-butyl hydroperoxide, and mercury) also blocked volume-stimulated taurine efflux, whereas efflux was unaffected by Ca2+ ionophore, phorbol ester, dibutyryl-adenosine 3',5'-cyclic monophosphate, vasopressin, or pretreatment with ouabain or furosemide. N-ethylmaleimide, diamide, 2,4-dinitrophenol, and iodoacetate plus KCN also inhibited the RVD. These findings suggest that, in contrast to hepatocytes from most vertebrate species, RVD in skate hepatocytes is associated with the release of only a small fraction of intracellular K+ but a substantial fraction of intracellular taurine and perhaps other organic osmolytes. This volume-activated taurine transport mechanism is energy and sulfhydryl group dependent and is not related to the taurine concentration gradient across the skate hepatocyte plasma membrane.

The volume-regulated anion channel is formed by LRRC8 heteromers – molecular identification and roles in membrane transport and physiology

2015 ◽  
Vol 396 (9-10) ◽  
pp. 975-990 ◽  
Author(s):  
Tobias Stauber
Keyword(s):  
Cell Biology ◽  
Regulatory Volume Decrease ◽  
Anion Channel ◽  
Cell Swelling ◽  
Transepithelial Transport ◽  
Organic Osmolytes ◽  
Physiological Processes ◽  
Starting Point ◽  
And Migration ◽  
Volume Decrease

Abstract Cellular volume regulation is fundamental for numerous physiological processes. The volume-regulated anion channel, VRAC, plays a crucial role in regulatory volume decrease. This channel, which is ubiquitously expressed in vertebrates, has been vastly characterized by electrophysiological means. It opens upon cell swelling and conducts chloride and arguably organic osmolytes. VRAC has been proposed to be critically involved in various cellular and organismal functions, including cell proliferation and migration, apoptosis, transepithelial transport, swelling-induced exocytosis and intercellular communication. It may also play a role in pathological states like cancer and ischemia. Despite many efforts, the molecular identity of VRAC had remained elusive for decades, until the recent discovery of heteromers of LRRC8A with other LRRC8 family members as an essential VRAC component. This identification marks a starting point for studies on the structure-function relation, for molecular biological investigations of its cell biology and for re-evaluating the physiological roles of VRAC. This review recapitulates the identification of LRRC8 heteromers as VRAC components, depicts the similarities between LRRC8 proteins and pannexins, and discussed whether VRAC conducts larger osmolytes. Furthermore, proposed physiological functions of VRAC and the present knowledge about the physiological significance of LRRC8 proteins are summarized and collated.

ATP regulates anion channel-mediated organic osmolyte release from cultured rat astrocytes via multiple Ca2+-sensitive mechanisms

2005 ◽  
Vol 288 (1) ◽  
pp. C204-C213 ◽  
Author(s):  
Alexander A. Mongin ◽  
Harold K. Kimelberg
Keyword(s):  
Intracellular Signaling ◽  
Anion Channel ◽  
Cell Swelling ◽  
Organic Osmolytes ◽  
Signaling Mechanism ◽  
Pharmacological Agents ◽  
Organic Osmolyte ◽  
Primary Astrocyte ◽  
Intracellular Ca ◽  
Rat Astrocytes

Ubiquitously expressed volume-regulated anion channels (VRACs) are activated in response to cell swelling but may also show limited activity in nonswollen cells. VRACs are permeable to inorganic anions and small organic osmolytes, including the amino acids aspartate, glutamate, and taurine. Several recent reports have demonstrated that neurotransmitters or hormones, such as ATP and vasopressin, induce or strongly potentiate astrocytic whole cell Cl− currents and amino acid release, which are inhibited by VRAC blockers. In the present study, we explored the intracellular signaling mechanisms mediating the effects of ATP on d-[3H]aspartate release via the putative VRAC pathway in rat primary astrocyte cultures. Cells were exposed to moderate (5%) or substantial (30%) reductions in medium osmolarity. ATP strongly potentiated d-[3H]aspartate release in both moderately swollen and substantially swollen cells. These ATP effects were blocked (≥80% inhibition) by intracellular Ca2+ chelation with BAPTA-AM, calmodulin inhibitors, or a combination of the inhibitors of protein kinase C (PKC) and calmodulin-dependent kinase II (CaMK II). In contrast, control d-[3H]aspartate release activated by the substantial hyposmotic swelling showed little (≤25% inhibition) sensitivity to the same pharmacological agents. These data indicate that ATP regulates VRAC activity via two separate Ca2+-sensitive signaling cascades involving PKC and CaMK II and that cell swelling per se activates VRACs via a separate Ca2+/calmodulin-independent signaling mechanism. Ca2+-dependent organic osmolyte release via VRACs may contribute to the physiological functions of these channels in the brain, including astrocyte-to-neuron intercellular communication.

Reactive oxygen species are important mediators of taurine release from skeletal muscle cells

2003 ◽  
Vol 284 (6) ◽  
pp. C1362-C1373 ◽  
Author(s):  
Niels Ørtenblad ◽  
Jette Feveile Young ◽  
Niels Oksbjerg ◽  
Jacob Holm Nielsen ◽  
Ian Henry Lambert
Keyword(s):  
Reactive Oxygen Species ◽  
Skeletal Muscle ◽  
Reactive Oxygen ◽  
Anion Channel ◽  
Muscle Cells ◽  
Skeletal Muscle Cells ◽  
Cell Swelling ◽  
Oxygen Species ◽  
Taurine Release ◽  
Taurine Efflux

The present study illustrates elements of the signal cascades involved in the activation of taurine efflux pathways in myotubes derived from skeletal muscle cells. Exposing primary skeletal muscle cells, loaded with 14C-taurine, to 1) hypotonic media, 2) the phospholipase A2 (PLA2) activator melittin, 3) anoxia, or 4) lysophosphatidyl choline (LPC) causes an increase in 14C-taurine release and a concomitant production of reactive oxygen species (ROS). The antioxidants butulated hydroxy toluene and vitamin E inhibit the taurine efflux after cell swelling, anoxia, and addition of LPC. The muscle cells possess two separate taurine efflux pathways, i.e., a swelling- and melittin-induced pathway that requires 5-lipoxygenase activity for activation and a LPC-induced pathway. The two pathways are distinguished by their opposing sensitivity toward the anion channel blocker DIDS and cholesterol. These data provide evidence for PLA2 products and ROS as key mediators of the signal cascade leading to taurine efflux in muscle.

Volume-sensitive taurine efflux from mammary tissue is not obliged to utilize volume-activated anion channels

1996 ◽  
Vol 16 (6) ◽  
pp. 459-465 ◽  
Author(s):  
D. B. Shennan ◽  
M. J. Cliff ◽  
P. Hawkins
Keyword(s):  
Anion Transport ◽  
Cell Swelling ◽  
Mammary Tissue ◽  
Anion Channels ◽  
Taurine Release ◽  
Amino Acid Transport System ◽  
Transport Inhibitors ◽  
Swelling Volume ◽  
Taurine Efflux ◽  
Stimulation Of

Cell-swelling, induced by a hyposmotic shock, activates the release of taurine from lactating rat mammary tissue expiants. The degree of stimulation of taurine efflux was dependent upon the extent of cell-swelling. Volume-sensitive taurine release was attenuated by the anion transport inhibitors NPPB, DIOA, DIDS, niflumate, flufenamate, mefenamate and diiodosalicylate but not by salicylate. Cell-swelling, following a hyposmotic challenge, did not increase the unidirectional efflux of radiolabelled I− or D-asparate from mammary tissue expiants. The results suggest that although mammary tissue expresses a volume-sensitive amino acid transport system which is inhibited by anion transport blockers the pathway has no identity with volume-activated anion channels.

Swelling-activated anion conductance in skate hepatocytes: regulation by cell Cl- and ATP

1996 ◽  
Vol 270 (1) ◽  
pp. C57-C66 ◽  
Author(s):  
P. S. Jackson ◽  
K. Churchwell ◽  
N. Ballatori ◽  
J. L. Boyer ◽  
K. Strange
Keyword(s):  
Mammalian Cells ◽  
Anion Channel ◽  
Fold Increase ◽  
Cation Binding ◽  
Cell Swelling ◽  
Organic Osmolytes ◽  
Main Function ◽  
Anion Conductance ◽  
Cation Permeability ◽  
Current Activation

Cell swelling activates an outwardly rectifying anion conductance in mammalian cells. The channel responsible for this conductance mediates volume-regulatory efflux of organic osmolytes such as taurine. We observed a similar conductance in hepatocytes from the skate Raja erinacea. Whole cell Cl- conductance was increased > 100-fold by a 2-fold increase in hepatocyte volume. The conductance was outwardly rectifying and had a relative cation permeability of approximately 0.2. Cation permeability was increased by reductions in patch pipette CsCl concentration, suggesting that the channel pore contains saturable anion and cation binding sites with different anion and cation affinities. The conductance had a broad anion selectivity and a relative taurine permeability of 0.17. Activation of the conductance required intracellular ATP or a nonhydrolyzable ATP analogue. Elevation of intracellular Cl- from 20 to 155 mM reduced current activation while the rate and extent of cell swelling were unaffected. Reduction of intracellular Cl- concentration to 5-10 mM caused spontaneous current activation without cell swelling. These results suggest that increases in cell Cl- levels increase the volume set point of the channel. We propose that the main function of the outwardly rectifying anion channel is nonselective transport of organic solutes.

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