Ca2+-dependent K+ transport in the Ehrlich ascites tumor cell (original) (raw)
Related papers
Separate, Ca2+-activated K+ and Cl− transport pathways in Ehrlich ascites tumor cells
The Journal of Membrane Biology, 1986
The net loss of KC1 observed in Ehrlich ascites cells during regulatory volume decrease (RVD) following hypotonic exposure involves activation of separate conductive K + and CI-transport pathways. RVD is accelerated when a parallel K § transport pathway is provided by addition of gramicidin, indicating that the K + conductance is rate limiting. Addition of ionophore A23187 plus Ca 2+ also activates separate K + and CItransport pathways, resulting in a hyperpolarization of the cell membrane. A calculation shows that the K + and C1 conductance is increased 14-and 10-fold, respectively. Gramicidin fails to accelerate the A23187-induced cell shrinkage, indicating that the C1-conductance is rate limiting. An A23187-induced activation of 42K and 36C1 tracer fluxes is directly demonstrated, RVD and the A23187-induced cell shrinkage both are: (i) inhibited by quinine which blocks the CaZ+-activated K + channel. (ii) unaffected by substitution of NO2 or SCN for CI-, and (iii) inhibited by the anti-calmodulin drug pimozide. When the K + channel is blocked by quinine but bypassed by addition of gramicidin, the rate of cell shrinkage can be used to monitor the CI-conductance. The CI conductance is increased about 60-fold during RVD. The volume-induced activation of the CI-transport pathway is transient, with inactivation within about 10 min. The activation induced by ionophore A23187 in Ca:+-free media (probably by release of Ca 2+ from internal stores) is also transient, whereas the activation is persistent in Ca2+-containing media. In the latter case, addition of excess EGTA is followed by inactivation of the C1-transport pathway. These findings suggest that a transient increase in free cytosolic Ca 2+ may account for the transient activation of the CI-transport pathway. The activated anion transport pathway is unselective, carrying both CI-, Br-, NO2, and SCN . The anti-calmodulin drug pimozide blocks the volume-or A23187-induced C1-transport pathway and also blocks the activation of the K + transport pathway. This is demonstrated directly by 42K flux experiments and indirectly in media where the dominating anion (SCN-) has a high ground permeability. A comparison of the A23187-induced K + conductance estimated from 42K flux measurements at high external K +, and from net K + flux measurements suggests single-file behavior of the Ca2+-activated K + channel. The number of Ca2+-activated K + channels is estimated at about 100 per cell.
The Journal of Membrane Biology, 1984
Ehrlich ascites tumor cells resuspended in hypotonic medium initially swell as nearly perfect osmometers, but subsequently recover their volume within 5 to 10 min with an associated KCI loss. 1. The regulatory volume decrease was unaffected when nitrate was substituted for C1-, and was insensitive to bumetanide and DIDS. 2. Quinine, an inhibitor of the Ca 2 § activated K + pathway, blocked the volume recovery. 3. The hypotonic response was augmented by addition of the Ca 2 + ionophore A23187 in the presence of external Ca z+, and also by a sudden increase in external Ca z+. The volume response was accelerated at alkaline pH. 4. The anti-calmodulin drugs trifluoperazine, pimozide, flupentixol, and chlorpromazine blocked the volume response. 5. Depletion of intracellular Ca z + stores inhibited the regulatory volume decrease. 6. Consistent with the low conductive C1 -permeability of the cell membrane there was no change in cell volume or C1 content when the K + permeability was increased with valinomycin in isotonic medium. In contrast, addition of the Ca z+ ionophore A23187 in isotonic medium promoted CI-loss and cell shrinkage. During regulatory volume decrease valinomycin accelerated the net loss of KC1, indicating that the conductive C1-permeability was increased in parallel with and even more than the K + permeability. It is proposed that separate conductive K + and C1-channels are activated during regulatory volume decrease by reiease of Ca z+ from internal stores, and that the effect is mediated by calmodulin.
Mechanism of Ca2+-dependent selective rapid K+-transport induced by propranolol in red cells
The Journal of Membrane Biology, 1977
Passive Ca 2+ influx is gradually enhanced by 0.5 to 5mM propranolol in fresh and phosphate ester-depleted human red cells. In fresh cells the active Ca 2 + efflux tends to counteract Ca 2+ uptake. Membrane hyperpolarization, induced by the K § transport that accompanies Ca 2 + uptake, further enhances the rate of Ca 2 uptake. The dissociated, positively charged form of propranolol seems to be crucial in the increase of passive Ca 2+ influx caused by the drug. The effect can be attributed to the release of structural Ca 2 + from the membrane (lipids).
Ca2+-activated K+ channels in human red cells. Comparison of single-channel currents with ion fluxes
Biophysical Journal, 1984
Exposure of the inner surface of intact red cells or red cell ghosts to Ca2" evokes unitary currents that can be measured in cell-attached and cell-free membrane patches. The currents are preferentially carried by K+ (PK/ PNa 17) and show rectification. Increasing the Ca2" concentration from 0 to 5 IuM increases the probability of the open state of the channels parallel to the change of K+ permeability as observed in suspensions of red cell ghosts. Prolonged incubation of red cell ghosts in the absence of external K+ prevents the Ca2" from increasing K+ permeability. Similarly, the probability to find Ca2"-activated unitary currents in membrane patches is drastically reduced. These observations suggest that the Ca2"-induced changes of K+ permeability observed in red cell suspensions are causally related to the appearance of the unitary K+ currents. Attempts to determine the number of K+ channels per cell were made by comparing fluxes measured in suspensions of red cells with the unitary currents in membrane patches as determined under comparable ionic conditions. At 100 mM KC1 in the external medium, where no net movements of K+ occur, the time course of equilibration of 86Rb+ does not follow a single exponential. This indicates a heterogeneity of the response to Ca2" of the cells in the population. The data are compatible with the assumption that 25% of the cells respond with Pk = 33.2 x 10-'4cm3/s and 75% with Pk = 3.
AJP: Cell Physiology, 2009
Addition of H2O2 (0.5 mM) to Ehrlich ascites tumor cells under isotonic conditions results in a substantial (22 ± 1%) reduction in cell volume within 25 min. The cell shrinkage is paralleled by net loss of K+, which was significant within 8 min, whereas no concomitant increase in the K+ or Cl− conductances could be observed. The H2O2-induced cell shrinkage was unaffected by the presence of clofilium and clotrimazole, which blocks volume-sensitive and Ca2+-activated K+ channels, respectively, and is unaffected by a raise in extracellular K+ concentration to a value that eliminates the electrochemical driving force for K+. On the other hand, the H2O2-induced cell shrinkage was impaired in the presence of the KCl cotransport inhibitor (dihydro-indenyl)oxyalkanoic acid (DIOA), following substitution of NO3− for Cl−, and when the driving force for KCl cotransport was omitted. It is suggested that H2O2 activates electroneutral KCl cotransport in Ehrlich ascites tumor cells and not K+ and ...
Ca2+-activated K+ permeability in human erythrocytes: Modulation of single-channel events
European Biophysics Journal, 1985
Elevated levels of intracellular Ca :÷ activate a K÷-selective permeability in the membrane of human erythrocytes. Currents through single channels were analysed in excised inside-out membrane patches. The effects of several ions that are known to inhibit K ÷ fluxes are described with respect to the single-channel events. The results suggest that the blocking ions can partly move into the channels (but cannot penetrate) and interact with other ions inside the pore. The reduction of single-channel conductance by Cs ÷, tetraethylammonium and Ba 2+ and of single-channel activity by quinine and Ba 2÷ is referred to different rates of access to the channel. The concentration-and voltage-dependent inhibition by ions with measurable permeability (Na ÷ and Rb ÷) can be explained by their lower permeability, with single-file movement and ionic interactions inside the pore.
Activation by high potassium of a novel voltage-operated Ca 2+ channel in rat spleen
British Journal of Pharmacology, 1997
1 High potassium produced a concentration-dependent contraction in rat isolated spleen. 2 The high potassium-induced contraction of rat spleen was abolished in Ca 2+ -free Krebs solution containing 1 mM EGTA, and the subsequent addition of 3 mM Ca 2+ restored the high potassiuminduced contraction to the control level. 3 Nifedipine, verapamil, diltiazem, Cd 2+ , Ni 2+ , Co 2+ , R-(+)-Bay K 8644 and pimozide inhibited and relaxed high potassium-induced contraction of rat spleen with IC 50 and EC 50 values much higher than those values in rat aorta. 4 In addition, high potassium-stimulated contraction of rat spleen was insensitive to o-conotoxin GVIA, o-conotoxin MVIIC and o-agatoxin IVA. 5 The high potassium-induced contraction of rat spleen was also unaected by tetrodotoxin (TTX), prazosin, chloroethylclonidine (CEC), yohimbine, propranolol, atropine, diphenhydramine, cimetidine, ketanserin, 3-tropanyl-indole-3-carboxylate, saralasin, indomethacin, nordihydroguaiaretic acid, GR32191B, domperidone, naloxone, chlorpromazine, suramin, (+)-2-amino-5-phosphonopentanoic acid, 6,7-dinitroquinoxaline-2,3-dione (DNQX), 877,606, lorglumide, PD 135, benextramine, amiloride, dantrolene, econazole, staurosporine and neomycin. 6 Forskolin and sodium nitroprusside relaxed high potassium-induced contraction of rat spleen with EC 50 values of 0.55+0.04 and 20.0+2.7 mM, respectively. 7 It is concluded that high potassium may activate a novel, pharmacologically uncharacterized voltageoperated Ca 2+ channel in rat spleen.