Swelling, NEM, and A23187 activate Cl(-)-dependent K+ transport in high-K+ sheep red cells

1987 ◽  
Vol 252 (2) ◽  
pp. C197-C204 ◽  
Author(s):  
H. Fujise ◽  
P. K. Lauf
Keyword(s):  
Metal Ion ◽  
Red Cells ◽  
Common Mechanism ◽  
Ionophore A23187 ◽  
Sheep Red Cells ◽  
High K ◽  
Sh Group ◽  
Low K ◽  
K Transport ◽  
Significant Activation

In low K+ (LK) sheep red cells a significant fraction of the total ouabain-resistant (OR) K+ flux is inhibited when Cl- is replaced by other anions of the Hofmeister series except Br- (Cl(-)-dependent K+ flux). In contrast, high K+ (HK) sheep red cells in isosmotic media did not possess any significant OR Cl(-)-dependent K+ flux when Cl- was replaced by NO3- or I-. However, exposure to hyposmotic solutions, treatment with the sulfhydryl (SH) group reagent N-ethylmaleimide (NEM) or with the bivalent metal ion (Me2+) ionophore A23187 in absence of external Me2+ caused a significant activation of Cl(-)-dependent K+ transport as measured with Rb+ as K+ congener. There was no Cl(-)-dependent Rb+ flux in A23187-treated cells when Mn2+, Mg2+, and Ca2+ were present at 1 mM concentrations, suggesting that cellular accumulation of these Me2+ is inhibitory. Similar to LK red cells, HK red cells failed to respond to A23187 when pretreated with NEM supporting the hypothesis proposed recently (Lauf, P. K. J. Membr. Biol. 88: 1-13, 1985) of a common mechanism of Cl(-)-dependent K+ transport activation. The magnitudes of the Cl(-)-dependent Rb+ fluxes in HK cells were much smaller than those elicited by identical treatments in LK red cells, and the effect of all interventions was not due to the presence of reticulocytes known to possess Cl(-)-dependent K+ transport.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Passive potassium transport in LK sheep red cells. Effects of anti-L antibody and intracellular potassium.

1976 ◽  
Vol 68 (6) ◽  
pp. 567-581 ◽  
Author(s):  
P B Dunham
Keyword(s):  
Maximum Velocity ◽  
Red Cells ◽  
Na Transport ◽  
Sheep Red Cells ◽  
The Common ◽  
K Concentration ◽  
Mode Of Transport ◽  
Low K ◽  
K Transport ◽  
External K

The passive K influx in low K(LK) red blood cells of sheep saturates with increasing external K concentration, indicating that this mode of transport is mediated by membrane-associated sites. The passive K influx, iMLK, is inhibited by external Na. Isoimmune anti-L serum, known to stimulate active K transport in LK sheep red cells, inhibits iMLK about twofold. iMLK is affected by changes in intracellular K concentration, [K]i, in a complex fashion: increasing [K]i from near zero stimulates iMLK, while further increases in [K]i, above 3 mmol/liter cells, inhibit iMLK. The passive K influx is not mediated by K-K exchange diffusion. The effects of anti-L antibody and [K]i on passive cation transport are specific for K: neither factor affects passive Na transport. The common characteristics of passive and active K influx suggest that iMLK is mediated by inactive Na-K pump sites, and that the inability to translocate Na characterizes the inactive pumps. Anti-L antibody stimulates the K pump in reticulocytes of LK sheep. However, anti-L has no effect on iMLK in these cells, apparently because reticulocytes do not have the inactive pump sites which, in mature LK cells, are a consequence of the process of maturation of circulating LK cells. The results also indicate that anti-L alters the maximum velocity of both active and passive K fluxes by converting pumps sites from a form mediating passive K influx to an actively transporting form.

A Nonselective Cation Permeabilization Pathway (Pcat) of Human Red Blood Cells (RBCs) Activated by Ca2+ and by Isotonic Dehydration.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1575-1575
Author(s):  
Robert M. Bookchin ◽  
Teresa Tiffert ◽  
Adaeze Muoma ◽  
Lynn Macdonald ◽  
Zipora Etzion ◽  
...  
Keyword(s):  
Terminal Condition ◽  
Atp Depletion ◽  
Common Mechanism ◽  
Ionophore A23187 ◽  
Transport Pathway ◽  
Experimental Conditions ◽  
Human Red Blood Cells ◽  
High K ◽  
Low K

Abstract In the course of experiments designed to study the patterns and distribution of dehydration rates of RBCs permeabilized to K+, we made the surprising observation that isotonic dehydration of RBCs elicits a spontaneous rehydration process by activating a non-selective cation permeability pathway which is highly stimulated by elevated [Ca2+]i. RBCs suspended in plasma-like, high-Na+, low-K+ buffers at 37°C and uniformly permeabilized to K+ either with valinomycin or by maximal Gardos channel activation (with ionophore A23187 + Ca2+) sustained net loss of KCl and water. The dehydration was maximal in 4–6 min when 10 mM SCN− replaced 10 mM Cl− to avoid the anion permeability rate limitation. The extent and distribution of RBC hydration was followed in time by the changes in osmotic lysis curves or by flow cytometry (Advia 120 hematology system). In all experimental conditions tested (varying Hct (<20%), buffer, dehydration rate), dehydration was followed by spontaneous rehydration, seen as a slow, progressive right shift in the hemolysis curves and volume distributions, whose patterns suggested substantial heterogeneity in RBC rehydration rates. By 20 hrs post-dehydration, rehydration had led to about 30% lysis, suggesting that in many RBCs, Na influx through Pcat exceeded its extrusion by the Na pump. Flame photometry showed progressive RBC Na gain during rehydration. Isotonic replacement of external Na by sucrose completely prevented rehydration, but choline or NMG only slowed it by about 50%, indicating that Pcat is poorly selective among organic and inorganic cations. Rehydration after valinomycin was much slower than in Ca2+-loaded cells, indicating powerful stimulation by Ca2+. Elevated [Ca2+]i also activated Pcat in RBCs equilibrated in high-K media, without prior dehydration. After inhibiting the Gardos channels (with charybdotoxin, clotrimazole or by extracting [Ca2+]i with excess EGTA), rehydration in high-K+ media occurred at a rate similar to that in high-Na+. Of the variety of transport-inhibitors tested so far on Pcat (furosemide, 0.5 mM; amiloride, 1 mM; TTX, 10μM; bumetanide, 1 mM; vanadate, 1 mM; quinine, 1 mM), only quinine was inhibitory (50%). Vanadate, which inhibits the Ca-pump and prevents the ATP depletion induced by Ca2+ loads, was stimulatory (125%) in the Ca2+-loaded RBCs, excluding ATP depletion as mediator of the Ca2+ effects. RBC aging is associated with increased cell density, reflecting isotonic RBC dehydration in vivo. Light, high-Na+ RBCs are found in small numbers in normal blood, and in greater numbers in sickle cell anemia and thalassemic blood. The observed response of RBCs to isotonic dehydration and increased [Ca2+]i, with activation of a transport pathway capable of generating swollen, high-Na+, low-K+ RBCs, raises the possibility that Pcat may represent the common mechanism leading from RBC dehydration of normally aged or pathologically dehydrated RBCs to a pre-lytic terminal condition of senescent RBCs.

Effect of N-ethylmaleimide on K transport in density-separated human red blood cells

1987 ◽  
Vol 253 (1) ◽  
pp. C7-C12 ◽  
Author(s):  
L. R. Berkowitz ◽  
D. Walstad ◽  
E. P. Orringer
Keyword(s):  
Red Blood Cells ◽  
Blood Cells ◽  
Red Cells ◽  
Sheep Red Cells ◽  
Human Red Blood Cells ◽  
High K ◽  
K Transport ◽  
Human Red Cells

N-ethylmaleimide (NEM) is a sulfhydryl-reacting agent known to stimulate chloride-dependent K transport in a variety of red cells. In high K sheep red cells, NEM-induced K movements are greater in magnitude in young cells compared with old cells. We hypothesized that human red cells might respond to NEM like high K sheep red cells. To test this idea, cells of various age were exposed to 0.5 mM NEM. We found that, after a 4-h incubation, young cells lost 50% of cell K, compared with 10% K loss in older cells. K loss in all fractions was inhibited by chloride replacement or furosemide.

Ion and water shifts produced by ammonium chloride in high K and low K red cells

1966 ◽  
Vol 68 (1) ◽  
pp. 19-24 ◽  
Author(s):  
Simo Salminen ◽  
Vesa Manninen
Keyword(s):  
Ammonium Chloride ◽  
Red Cells ◽  
High K ◽  
Low K

Several cation transporters and volume regulation in high-K dog red blood cells

1991 ◽  
Vol 260 (3) ◽  
pp. C589-C597 ◽  
Author(s):  
H. Fujise ◽  
I. Yamada ◽  
M. Masuda ◽  
Y. Miyazawa ◽  
E. Ogawa ◽  
...  
Keyword(s):  
Red Blood Cells ◽  
Blood Cells ◽  
Volume Regulation ◽  
Cation Composition ◽  
Influx Rate ◽  
Physiological Conditions ◽  
High K ◽  
Dependent Transport ◽  
Low K ◽  
K Transport

Normal dog red blood cells lack the Na-K pump, and their cation composition is low K and high Na (LK). Recently, a dog was found with red blood cells containing high K and low Na concentrations (HK) due to the existence of the Na-K pump. In the present study, cation transport and volume regulation in HK cells were compared with those of LK cells. HK cells showed not only Rb influx through a Na-K pump, but also Rb influx through a Cl-dependent K transporter. The Rb influx rate through the Na-K pump was 0.65-1.44 mmol.l cells-1.h-1 in Cl and 1.75-2.24 mmol.l cells-1.h-1 in NO3, in HK cells, but only trace activities are found in LK cells. In HK cells, the Rb influx rate through Cl-dependent K transport was 0.36-0.96 mmol.l cells-1.h-1, and it was enhanced in swollen cells but vanished in shrunken cells. In LK cells, the transport was evident only in swollen cells. The original volume of swollen HK cells was restored by water extrusion promoted by Cl-dependent transport. The Na-Ca exchange transporter, which works as a volume regulator in LK cells, functioned in HK cells only when they were loaded with Na. Hence, the exchange transporter is latent in HK cells under physiological conditions. Moreover, the exchange transporter could restore the cell volume in swollen and Na-loaded HK cells. However, the volume in HK cells was still larger than that in LK cells, while the Na-Ca exchange transporter was working.(ABSTRACT TRUNCATED AT 250 WORDS)

Cl-dependent K transport in a pure population of volume-regulating human erythrocytes

1989 ◽  
Vol 256 (4) ◽  
pp. C858-C864 ◽  
Author(s):  
W. C. O'Neill
Keyword(s):  
Cell Volume ◽  
Regulatory Volume Decrease ◽  
Human Erythrocytes ◽  
Red Cells ◽  
Hypotonic Medium ◽  
K Cells ◽  
High K ◽  
The Relationship ◽  
Low K ◽  
Volume Decrease

Swelling of human red cells activates a putative K-Cl cotransport that is not present at normal cell volume and that disappears after several hours. To determine whether regulatory volume decrease (RVD) is occurring in human erythrocytes and is responsible for the inactivation of K-Cl cotransport, the relationship between cell volume and the inactivation and reactivation of volume-sensitive (VS) K-Cl cotransport was studied. VS K influx into high K cells was transient, whereas influx into low K cells (prepared with nystatin), which are unable to shrink via K efflux, remained fully activated. Likewise, VS K efflux into hypotonic medium disappeared after 100 min in a low K medium but remained activated in a high K medium that prevented cell shrinkage. Cells that had been preincubated in hypotonic medium to inactivate VS K-Cl cotransport showed no significant recovery of VS cotransport after a 6-h incubation in isotonic medium but showed full restoration of VS cotransport after treatment with nystatin in isotonic medium to reequilibrate cell water. A pure fraction of volume-regulating (VR) cells was subsequently isolated by preincubating red cells in hypotonic medium and then subjecting them to further hypotonicity to lyse all non-VR cells. The 2.5% of cells that remained consisted of 16% reticulocytes and exhibited a Cl-dependent RVD in hypotonic medium. VS K-Cl cotransport was enriched 10-fold and Na-K-Cl cotransport was enriched 12-fold in these cells, whereas the enrichment of N-ethylmaleimide (NEM)-activated K-Cl cotransport was only threefold.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Active Sodium and Potassium Transport in High Potassium and Low Potassium Sheep Red Cells

1971 ◽  
Vol 58 (4) ◽  
pp. 438-466 ◽  
Author(s):  
P. G. Hoffman ◽  
D. C. Tosteson
Keyword(s):  
Cell Types ◽  
Red Cells ◽  
Kinetic Characteristics ◽  
High Potassium ◽  
Cation Composition ◽  
Sheep Red Cells ◽  
Pump Rate ◽  
Low Potassium ◽  
K Transport ◽  
External K

The kinetic characteristics of the ouabain-sensitive (Na + K) transport system (pump) of high potassium (HK) and low potassium (LK) sheep red cells have been investigated. In sodium medium, the curve relating pump rate to external K is sigmoid with half maximal stimulation (K1/2) occurring at 3 mM for both cell types, the maximum pump rate in HK cells being about four times that in LK cells. In sodium-free media, both HK and LK pumps are adequately described by the Michaelis-Menten equation, but the K1/2 for HK cells is 0.6 ± 0.1 mM K, while that for LK is 0.2 ± 0.05 mM K. When the internal Na and K content of the cells was varied by the PCMBS method, it was found that the pump rate of HK cells showed a gradual increase from zero at very low internal Na to a maximum when internal K was reduced to nearly zero (100% Na). In LK cells, on the other hand, no pump activity was detected if Na constituted less than 70% of the total (Na + K) in the cell. Increasing Na from 70 to nearly 100% of the internal cation composition, however, resulted in an exponential increase in pump rate in these cells to about ⅙ the maximum rate observed in HK cells. While changes in internal composition altered the pump rate at saturating concentrations of external K, it had no effect on the apparent affinity of the pumps for external K. These results lead us to conclude that the individual pump sites in the HK and LK sheep red cell membranes must be different. Moreover, we believe that these data contribute significantly to defining the types of mechanism which can account for the kinetic characteristics of (Na + K) transport in sheep red cells and perhaps in other systems.

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