42K Efflux Research Articles

K+-induced swelling of the dogfish shark (Squalus acanthias) rectal gland cells is associated with changes of the cytoskeleton

Dogfish shark (Squalus acanthias) rectal gland cells swell massively when incubated in elasmobranch media in which Na+ was equivalently replaced by K+; this swelling was abolished when the impermeant gluconate replaced Cl-, while the cell depolarization was comparable in both media. The K+-effect was associated with (a) an increase of the steady-state 42K (and 86Rb) efflux (particularly of the rate constant of the fast cellular efflux component) and a rearrangement of the respective cellular pools of K+; (b) an alteration of cell morphology and the pattern of the F-actin staining along the basolateral cell membrane as revealed with fluorescent analogs of phallacidin. These changes were independent of cell volume, being identical in KCl and K-gluconate media. The observations were specific for K+ (and Rb+): replacement of media Na+ by Li+ (which is not actively extruded by the cells), or the presence of ouabain, produced only minor swelling without affecting cell morphology and F-acting distribution. The results are consistent with the following view: as opposed to Na+ or Li+ media, the K+-induced changes of the cortical F-actin component of the cytoskeleton permit the observed massive cell swelling due to the osmotic contribution of intracellular impermeant anion(s).

Effects of aging on agonist-activated 86Rb efflux in arteries of Fischer 344 rats.

Segments of thoracic aorta (DTA), tail artery (TA), and mesenteric artery branches (MAB) were obtained from male Fischer 344 rats at ages of 1, 2, 6, 12, 24, and 30 mo and were used to determine the effects of aging on agonist-activated 86Rb (and 42K) efflux. At all three arterial sites, basal efflux decreased during development (1-6 mo), but no further changes were observed with aging (6-30 mo). The initial efflux response to 10 microM norepinephrine (NE) in the presence of 1 microM propranolol exhibited either no change (DTA) or an increase (TA and MAB) during development (1-6 mo), but all three sites showed a large decrease during aging (6-30 mo). Changes in the steady-state response to NE paralleled changes in the basal efflux at all ages and arterial sites. The initial efflux response to 75 mM K+-physiological salt solution (PSS) for the DTA in the presence of 1 microM phentolamine and 1 microM propranolol decreased during development followed by an increase during aging, whereas for the TA and MAB, there were no significant changes with age. The steady-state efflux response to K+ decreased during development at all three sites but was increased only for the DTA during aging. The steady-state efflux response to K+ was not altered for the TA and MAB during aging. Efflux responses using 42K were qualitatively similar, but rate constants were quantitatively larger than those with 86Rb at all three arterial sites and at all ages.(ABSTRACT TRUNCATED AT 250 WORDS)

K+ transport and membrane potentials in isolated rat parotid acini.

42K+ transport properties of isolated rat parotid acini were characterized concomitant with measurements of membrane potentials (Em) by means of the fluorescent dye diSC3-(5). In unstimulated acini suspended in a 5 mM K+ buffer, Em was governed by the K+ and Cl- gradients and amounted to about -59 mV, a value that remained unaffected on cholinergic stimulation. In unstimulated acini, 42K+ influx was largely mediated by the Na+-K+ pump, and the residual influxes were mediated by a bumetanide-sensitive component (cotransport system) and by K+ channels. Efflux of 42K+ was largely mediated by a bumetanide-sensitive component and by K+ channels. In the unstimulated state, the cotransport system was mediating K+-K+ exchange without contributing to the net uptake of K+. Within 10 s after stimulation, a approximately 10-fold increase in the acinar K+ conductance (gK) occurred, resulting in a rapid net efflux of K+ that amounted to approximately 3.8 mmol.l cells-1.s-1. Measurements of 42K+ fluxes as a function of the external K+ concentration revealed that in the stimulated state gK increases when external K+ is raised from 0.7 to 10 mM, consistent with an activation of acinar gK by the binding of external K+ to the channel. 42K+ flux ratios as well as the effect of the K+ channel inhibitor from scorpion venom (LQV) suggest that approximately 90% of K+ transport in the stimulated state is mediated by "maxi" K+ channels.

The potassium channel opening action of pinacidil; studies using biochemical, ion flux and microelectrode techniques.

In rat aorta and rat portal vein, (-)- and (+)-pinacidil each produced a concentration-dependent inhibition of tension development. Although the (-) isomer was the more potent, concentration effect curves for each isomer were steep with similar slopes. In rat portal vein, tetraethylammonium and procaine antagonised the relaxant effect of (+/-)-pinacidil, whereas 3,4-diamino-pyridine was without effect. Intracellular microelectrode recording in rat portal vein showed that low concentrations of (+/-)-pinacidil reduced the duration of multispike electrical complexes. In both rat aorta and rat portal vein, higher concentrations of (+/-)-pinacidil hyperpolarised the membrane towards the potassium equilibrium potential. (+/-)-Pinacidil increased 86Rb efflux from rat aorta and rat portal vein in a concentration dependent manner. In a separate study, (+/-)-pinacidil increased 42K efflux from rat portal vein. (+/-)-Pinacidil had no effect on cyclic GMP or cyclic AMP levels in rat aorta. It is concluded that pinacidil opens 86Rb-permeable potassium channels in rat aorta and rat portal vein. This mechanism is independent of cyclic nucleotide changes and may be responsible for the antihypertensive effect of pinacidil.

Altered chloride transport in arteries from aldosterone salt-hypertensive rats

Chronic infusion of d-aldosterone into uninephrectomized rats on high sodium intake resulted in a significant increase in systolic blood pressure. The steady state efflux of 36Cl in aorta and femoral artery of hypertensive rats was significantly elevated compared to that in arteries of control rats. These elevations in chloride efflux persisted in the presence of the Cl-HCO3 exchange inhibitor, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), and in Cl-HCO3 free solution. The Cl-HCO3-dependent component of the 36Cl efflux was also higher under some conditions. DIDS (100 mumol/l), also significantly reduced the increase in 36Cl efflux caused by norepinephrine (NE) in one of four groups, but had no significant effect on the NE stimulated 42K efflux. Contractile responses to both NE and KCl were unaffected by DIDS treatment. Therefore, it appears that Ca influx and release mechanisms function normally in the presence of DIDS. Similarly, DIDS had either a small or no effect on 42K movements under basal conditions. A slight reduction in the passive efflux of 24Na was observed in aortic smooth muscle exposed to DIDS with no significant effect on active Na transport. These findings indicate that DIDS is reasonably selective for Cl exchange sites in vascular smooth muscle during an exposure of 15 min or less and has little effect on cationic transport processes. The application of this blocker as well as anion substitution indicates that an elevation in the 36Cl leak efflux and related Cl permeability of arterial smooth muscle is associated with aldosterone-salt hypertension.

Altered biochemical and functional responses in aorta from hypertensive rats.

Factors that lead to supersensitivity of vascular smooth muscle to norepinephrine during aldosterone-salt-induced hypertension in rats appear to reside beyond ligand-alpha-adrenergic receptor binding, which we have shown previously to be normal. The objective of this study was to determine whether significant shifts occur in the coupling between receptors and the production of putative second messengers. Measures of [3H]myo-inositol phosphates in aorta (endothelium removed) exhibited a concentration-dependent increase to norepinephrine, with the 50% response shifted significantly to the left in the hypertensive group (7.0 +/- 0.9 X 10(-7) M in 8 control rats vs 1.1 +/- 0.2 X 10(-7) M in 8 hypertensive rats; p less than 0.001). The production of [32P]phosphatidic acid was also shifted (6.5 +/- 2.5 X 10(-7) M in 16 control vs 1.9 +/- 0.8 X 10(-7) M in 12 hypertensive rats; p less than 0.05). The functional responses of 42K efflux and contraction to norepinephrine were also significantly shifted threefold to 15-fold in the hypertensive group (p less than 0.001), but the 50% response typically occurred at a 10 to 100 times lower concentration than that for the production of myo-inositol phosphates and phosphatidic acid. The amplification between receptor occupancy and functional responses apparently occurs beyond the production of phosphoinositide metabolites. The fivefold shift in the 50% response of biochemical end points for the hypertensive group accounted for most of the shift (sixfold) in the functional end points.(ABSTRACT TRUNCATED AT 250 WORDS)

Alterations in cation homeostasis in cultured chick ventricular cells during and after recovery from adenosine triphosphate depletion.

Alterations in cation homeostasis during and after recovery from myocardial ischemia may account for some of the reversible and irreversible components of myocardial cell injury. To investigate possible mechanisms involved, we exposed cultured layers of spontaneously contracting chick embryo ventricular cells to media containing 1 mM cyanide (CN) and 20 mM 2-deoxyglucose (2-DG), and zero glucose for up to 6 h, and then allowed cultured cells to recover in serum-free culture medium for 24 h. Changes in Na, K, and Ca contents, 42K uptake and efflux, ATP content, cell water content, and lactate dehydrogenase (LDH) release were measured, and compared with changes produced by exposure to 10(-3) M ouabain and severe hypoxia. Exposure to CN and 2-DG caused marked increase in cell Na (sevenfold) and Ca (fivefold) contents, and a decrease in K content (one-fifth normal), coincident with ATP depletion to one-tenth normal levels. This produced only slight cell injury, evidenced by increased LDH release. Recovery for 24 h resulted in return to near normal values (expressed in nanomoles per milligram of protein) of Na, Ca, and ATP contents. However, there was failure of cell K content to return to normal, associated with a persistent reduced net uptake of 42K, and an increase in the rate of 42K efflux. These abnormalities in K homeostasis were associated with a decrease in cell volume and water content per milligram of protein. More marked ATP depletion (to 1/100 normal values) was produced by hypoxia plus 2-DG and zero glucose, and was associated with much more severe cell injury manifested by LDH loss. Ouabain exposure resulted in a much greater Ca gain (20-30-fold), relative to increase in Na content, than did either CN and 2-DG or hypoxia; and ouabain effects were not reversible (after a 15-fold or greater increase in Ca content was produced) and were associated with significant LDH release. We conclude that these cells are resistant to cell injury caused by moderately severe Ca overload and ATP depletion produced by exposure to CN and 2-DG. However, metabolic inhibition of ATP production produces persistent abnormalities in K homeostasis, associated with functional abnormalities.

Vasodilatation with pinacidil. Mode of action in rat resistance vessels.

Pinacidil is a newly developed antihypertensive vasodilator, proposed to belong to the new group of smooth muscle relaxants, the K+ channel openers. The in vitro effects of pinacidil on induced tone, smooth muscle membrane potential and 86Rb and 42K efflux from rat resistance vessels (internal diameter about 200 microns) were studied. Tone induced with noradrenaline was concentration-dependently inhibited by pinacidil. Responses to electrical field stimulation were also inhibited. However, tone induced with high K+ depolarisation, noradrenaline in the presence of high K+, caffeine-induced contractions and noradrenaline contractions in the presence of felodipine were little affected by pinacidil. Pinacidil caused concentration-dependent hyperpolarisation of the resting smooth muscle. Pinacidil caused only a small and transient increase of the 86Rb efflux rate constant, while the same concentrations of pinacidil produced a significant increase in the 42K efflux rate constant. Our results seem to indicate that the relaxant effect of pinacidil is the result of an increase in K+ permeability, thus causing hyperpolarisation and relaxation. The opened K+ channels appear to be selective for K+ over Rb+.

Dissociation of actions of BRL 34915 in the rat portal vein

In rat portal vein, 0.5 and 5.0 μM BRL 34915 [(±)-6-cyano-3,4-dihydro-2,2-dimethyl-trans-4-(2-oxo-l-pyrrolidyl)-2H-benzo[b]pyran-3-ol] abolished spontaneous rhythmic movements and norepinephrine (NE)-induced tension responses, respectively. Only the higher (5 μM) concentration of BRL 34915 increased 42K efflux and inhibited the NE-induced increase in 42K efflux. These results suggest that BRL 34915 inhibits spontaneous rhytmic movements and NE-induced tension responses by differing mechanisms of action and that only the block of the NE-induced tension response is related to K + permeability or conductance changes.

Comparison of simultaneous electrophysiologic and mechanical effects of verapamil and nifedipine in cultured chick embryo ventricular cells

In order to examine possible differences in mechanisms of action of nifedipine and verapamil, simultaneous recordings of membrane potential and cell motion were made during exposure of cultured chick embryo ventricular cells to these drugs. Both verapamil and nifedipine produced a rapid negative inotropic effect and excitation-contraction coupling delay presumably related to the ability of both agents to inhibit Ca2+ influx via the slow Ca channel, and thus decrease the [Ca2+]i transient. However, for an equivalent negative inotropic effect, verapamil produced relatively more membrane diastolic depolarization and slowing of repolarization than did nifedipine. Verapamil also produced a more marked decrease in +dV/dtmax. In ion flux studies, it was demonstrated that verapamil produced a significant inhibition of 24Na influx, relative to control, whereas nifedipine did not. Verapamil also produced a slight decrease in 42K efflux, of borderline significance. These findings support previous results which indicate there are differences in the mechanisms of action of these Ca2+-channel blocking drugs.

The role of extracellular sodium on heart muscle energetics.

A study has been made of changing external sodium concentration [Na]e, over the range 75 to 200 mmol X l-1, on contractile parameters and heat production in isolated, arterially perfused, interventricular rabbit septa.- The observed changes in maximum rate of contraction with [Na]e, either in the presence of a constant external Ca concentration [Ca]e or in the presence of a constant [Na]e2/[Ca]e ratio, paralleled those observed for tension development (T). On the other hand the maximal rate of relaxation (-Tmax) and the ratio -Tmax/T increased. While the ratio between active heat production and developed tension remained unaltered (0.111 +/- 0.003 mJ X mN-1 X g-1 dry weight), resting heat production increased with [Na]e2 with a slope of 95 +/- 18 mW X g-1 X mol-2 X l2. Under resting conditions, a decrease in [Na]e of 50 mmol X l-1 induced a fall in 42K uptake of about 16 nmol X s-1 X g-1 without changes in 42K efflux, suggesting that such an intervention depresses K influx. If the depressed K influx, induced by a decrease in [Na]e of 50 mmol X l-1, is associated with a decrease in Na-K pump activity, a fall in resting heat production of about 0.64 mW X g-1 would be expected. This represent 56% of the calculated change in the resting heat production, 1.14 +/- 0.22 mW X g-1 (mean +/- one confidence interval), suggesting that some process in addition to a depressed Na-K pump activity may be altered by changes in [Na]e.

Voltage dependence of the Ca2+-activated K+ conductance of human red cell membranes is strongly dependent on the extracellular K+ concentration.

The conductance of the Ca2+-activated K+ channel (gK(Ca)) of the human red cell membrane was studied as a function of membrane potential (Vm) and extracellular K+ concentration ([K+]ex). ATP-depleted cells, with fixed values of cellular K+ (145 mM) and pH (approximately 7.1), and preloaded with approximately 27 microM ionized Ca were transferred, with open K+ channels, to buffer-free salt solutions with given K+ concentrations. Outward-current conductances were calculated from initial net effluxes of K+, corresponding Vm, monitored by CCCP-mediated electrochemical equilibration of protons between a buffer-free extracellular and the heavily buffered cellular phases, and Nernst equilibrium potentials of K ions (EK) determined at the peak of hyperpolarization. Zero-current conductances were calculated from unidirectional effluxes of 42K at (Vm-EK) approximately equal to 0, using a single-file flux ratio exponent of 2.7. Within a [K+]ex range of 5.5 to 60 mM and at (Vm-EK) greater than or equal to 20 mV a basic conductance, which was independent of [K+]ex, was found. It had a small voltage dependence, varying linearly from 45 to 70 microS/cm2 between 0 and -100 mV. As (Vm-EK) decreased from 20 towards zero mV gK(Ca) increased hyperbolically from the basic value towards a zero-current value of 165 microS/cm2. The zero-current conductance was not significantly dependent on [K+]ex (30 to 156 mM) corresponding to Vm (-50 mV to 0). A further increase in gK(Ca) symmetrically around EK is suggested as (Vm-EK) becomes positive. Increasing the extracellular K+ concentration from zero and up to approximately 3 mM resulted in an increase in gK(Ca) from approximately 50 to approximately 70 microS/cm2. Since the driving force (Vm-EK) was larger than 20 mV within this range of [K+]ex this was probably a specific K+ activation of gK(Ca). The Ca2+-activated K+ channel of the human red cell membrane is an inward rectifier showing the characteristic voltage dependence of this type of channel.

Effect of channel blockers on potassium efflux from metabolically exhausted frog skeletal muscle.

1. 86Rb and 42K have been used to assess potassium exchange in frog skeletal muscle which had been metabolically exhausted by electrical stimulation (1 Hz) after treatment with 2 mM-cyanide and 1 mM-iodoacetate. These conditions led to the development of rigor. 2. Poisoning by itself induced a small but variable increase in tracer efflux. Complete mechanical exhaustion subsequent to electrical stimulation was, however, accompanied by a 5-6 fold increase in the rate coefficient for both 86Rb and 42K efflux. In the case of rubidium this was maintained for at least 20 min and often for up to 1 h. 3. The increase in tracer efflux induced by metabolic exhaustion was inhibited by barium (0.03-5 mM) in a reversible and concentration-dependent manner. Inhibition was also observed with glibenclamide (3-100 microM), tolbutamide (0.3-2 mM), TEA (5-100 mM) and the local anaesthetics lignocaine (1-3 mM) and tetracaine (1 mM). Quinine produced a dual response consisting of an inhibitory component which was most clearly seen at low concentrations (0.3 mM) and an enhancement of tracer efflux that became increasingly dominant at higher concentrations (1-10 mM). 4. Both apamin (30 and 100 nM) and Israeli scorpion (Leiurus quinquestriatus) venom (16 micrograms ml-1) produced little or no block of the tracer efflux activated by metabolic exhaustion. Similarly 4-aminopyridine (3 mM) and decamethonium (0.3 mM) were without obvious effect. 5. It is concluded that metabolic exhaustion of frog skeletal muscle leads to an increased permeability to both 42K and 86Rb. Our results with channel blockers suggest that this K+ permeability can be attributed neither to the delayed rectifier nor to an apamin- or charybdotoxin-sensitive calcium-activated K+ permeability (PK(Ca) but may be predominantly due to activation of ATP-sensitive channels similar to those found in the beta-cells of pancreatic islets.

Influence of external barium and potassium on potassium efflux in depolarized frog sartorius muscles.

Efflux of 42K+ was measured in frog sartorius muscles equilibrated in depolarizing solutions with external K+ concentrations [( K+]0) between 75 and 300 mM and NaCl concentrations of 60, 120, or 240 mM. For several combinations of KCl and NaCl, steady-state internal potentials (Vi) were the same for different [K+]0. For the range of Vi examined, K+ efflux occurs principally through the K+ inward rectifier channels. When external K+ is removed Vi remains constant for 2 to 3 hr because of the high membrane conductance to Cl-, but K+ efflux drops by about one order of magnitude. External Ba2+ in the presence or absence of external K+ produces an inhibition of K+ efflux described by a relation of the form u = (u1/(1 + C [Ba2+]0] + u2, where u is the uninhibited fraction of K+ efflux; u1, u2 and C are constants; and u1 + u2 = 1. C depends both on [K+]0 and Vi. When [K+]0 greater than or equal to 75 mM, increasing [K+]0 at constant Vi reduces Ba2+ sensitivity. For constant Vi greater than or equal to -30 mV, Ba2+ sensitivity is less when [K+]0 = 0 than when [K+]0 greater than or equal to 75 mM. When [K+]0 = 0, Ba2+ sensitivity decreases as Vi is made more positive. The dependence of the Ba2+ sensitivity on Vi at constant [K+]0 is greater when [K+]0 = 0 than when [K+]0 greater than or equal to 75 mM. Both the activation of K+ efflux by external K+ and the Ba2+ inhibition of K+ efflux can be explained on the basis of two membrane control sites associated with each channel. When both sites are occupied by K+, the channels are in a high flux state. When one or both sites are empty, the channels are in a low, nonzero flux state. When Ba2+ occupies either site, K+ efflux is further reduced. The reduction of Ba2+-sensitivity by increasing [K+]0 at high [K+]0 is attributable to the displacement of Ba2+ from the control sites by K+. The increased Ba2+ sensitivity produced by going from [K+]0 = 0 to [K+] greater than or equal to 75 mM when Vi greater than or equal to -30 mV is attributable to states in which Ba2+ occupies one site and K+ the other when [K+]0 not equal to 0.(ABSTRACT TRUNCATED AT 400 WORDS)

Zinc inhibition of potassium efflux in depolarized frog muscle and its modification by external hydrogen ions and diethylpyrocarbonate treatment.

Efflux of 42K+ was measured in frog sartorius muscles equilibrated in hyperosmotic depolarizing solutions. At the internal potentials obtained, K+ passes mainly through the inward rectifier potassium channels. Inhibition of K+ efflux by external Zn2+ (0.25 to 15 mM) differs in three significant ways from inhibition by Ba2+. (1) The dose-response relation does not correspond to action at a single site. (2) The Zn2+-sensitivity of K+ efflux does not depend on [K+]0 at constant internal potential. (3) Zn2+ inhibition is reduced by hydrogen ions, while Ba2+ inhibition is unaffected. Further, the Ba2+-sensitivity of K+ efflux is not altered by a half-inhibiting Zn2+ concentration, suggesting that the two ions do not interact at a common site. The histidine-modifying reagent diethylpyrocarbonate (DEPC) reduces Zn2+ inhibition. After DEPC treatment Zn2+ inhibition is further reduced by low pH. DEPC has little effect on Ba2+ inhibition. Zn2+ inhibition is not altered by treatment with the sulfhydryl reagents 5,5'-dithio-bis(2-nitrobenzoic acid) or dithiothreitol. The results can be described by either of two models in which two sites can bind Zn2+ and one or both of the sites may also bind H+. When both sites bind Zn2+, K+ efflux is inhibited, and a third site may then bind H+. The effects of DEPC can be accounted for by a decrease in H+ affinity of the first two sites by a factor of 50, and a decrease in Zn2+ affinity of these sites and of the H+ affinity of the third site by about one order of magnitude.

Potassium and rubidium permeability and potassium conductance of the beta-cell membrane in mouse islets of Langerhans.

The efflux of 42K+ and 86Rb+ has been studied in collagenase-isolated normal mouse islets of Langerhans. In double-labelled experiments, the ratio of efflux rate constants of Rb+ and K+ (kRb/kK) was 0.80 in 5 mM-K+ and 0 glucose. The ratio was unaffected by glucose concentrations up to 22.2 mM. In the presence of 50 mM-K+, 0 glucose, the ratio increased to 0.91 and in 50 mM-Rb+, 0 glucose and 0 K+, the ratio was 1.06. With these limitations, the results indicate that 86Rb+ is an acceptable isotope for K+. Using the Goldman model, K+ and Rb+ permeability coefficients and K+ slope and chord conductances were computed. The permeabilities decreased in glucose and in high K+ or high Rb+. In the case of high external K+, the K+ conductances increased. Also there may be more than one type of K+ channel with differing selectivities to K+ and Rb+. The addition of glucose in the presence of 50 mM-K+ had no further effect on Rb+ permeability. It is suggested that there are about ten small K+ channels open in the resting beta-cell and that progressive closure of these channels is involved in the depolarization of the cell membrane that initiates spike activity.

Potassium loss from rabbit myocardium during hypoxia: Evidence for passive efflux linked to anion extrusion

To determine the effects of permeant and impermeant anions and of osmolarity on potassium (K+) exchange, the net uptake and efflux of 42K+ were recorded in the isolated arterially perfused rabbit septum. Perfusion with solution made hyperosmolar by adding NaCl (30 mM) or sucrose (60 mM) caused similar increases of 42K+ uptake which were reversible on returning to the control solution. Washout experiments showed that the loss of K+ on returning to the control perfusate was due to a decreased influx probably mediated by inhibition of the sodium pump. The effects of anions were studied by replacing chloride in the control solution with the inert and impermeant substitute isethionate (114 mM) or by loading the myocardium with sodium dimethyloxazeolidinedione (NaDMO, 30 mM) under isosmotic condition and switching to a perfusate containing sodium isethionate (30 mM). In both these conditions a reduction of 42K content could be detected and was attributable to an increased efflux. During hypoxic substrate free perfusion K+ loss was due to an increased efflux with no evidence for altered influx of potassium. The extrusion of accumulated anions from the myocardium could be the major determinant of the early potassium loss during hypoxia and ischaemia.

Na+ and K+ transport at basolateral membranes of epithelial cells. II. K+ efflux and stoichiometry of the Na,K-ATPase.

Changes of 42K efflux (J23K) caused by ouabain and/or furosemide were measured in isolated epithelia of frog skin. From the kinetics of 42K influx (J32K) studied first over 8-9 h, K+ appeared to be distributed into readily and poorly exchangeable cellular pools of K+. The readily exchangeable pool of K+ was increased by amiloride and decreased by ouabain and/or K+-free extracellular Ringer solution. 42K efflux studies were carried out with tissues shortcircuited in chambers. Ouabain caused an immediate (less than 1 min) increase of the 42K efflux to approximately 174% of control in tissues incubated either in SO4-Ringer solution or in Cl-Ringer solution containing furosemide. Whereas furosemide had no effect on J23K in control tissues bathed in Cl-rich or Cl-free solutions, ouabain induced a furosemide-inhibitable and time-dependent increase of a neutral Cl-dependent component of the J23K. Electroconductive K+ transport occurred via a single-filing K+ channel with an n' of 2.9 K+ efflux before ouabain, normalized to post-ouabain (+/- furosemide) values of short-circuit current, averaged 8-10 microA/cm2. In agreement with the conclusions of the preceding article, the macroscopic stoichiometry of ouabain-inhibitable Na+/K+ exchange by the pump was variable, ranging between 1.7 and 7.2. With increasing rates of transepithelial Na+ transport, pump-mediated K+ influx saturated, whereas Na+ efflux continued to increase with increases of pump current. In the usual range of transepithelial Na+ transport, regulation of Na+ transport occurs via changes of pump-mediated Na+ efflux, with no obligatory coupling to pump-mediated K+ influx.

Potassium efflux from single skinned skeletal muscle fibers

The efflux of 42K from single, skinned (sarcolemma removed) skeletal muscle fibers has been determined. Isotope washout curves are kinetically complex and can be fit as the sum of three exponentials, including a fast component (k = 0.25 s-1) with a pool size equivalent to 91% of the fiber volume, an intermediate component (k = 0.08 s-1) equivalent to 6% of the fiber volume, and a slow component (k = 0.008 s-1) equivalent to 0.5% of fiber volume. Only the intermediate kinetic component is significantly affected by pretreatment of fibers with detergent. Efflux curves from detergent-treated fibers could be fit as the sum of two exponentials with coefficients and rate constants comparable to those of the fast and slow component of washout of untreated controls. Similarly the washout of [14C]sucrose can be described as the sum of two exponentials. We conclude that the intermediate component of 42K washout results from the movement of ions from a membrane bound space within the skinned fiber. Because of its relative volume, the sarcoplasmic reticulum seems to be a reasonable choice as a structural correlate for this component. Our estimate of the potassium permeability for the sarcoplasmic reticulum (SR) based on the efflux data is 10(-7) cm/s. This value is less than previous estimates from isolated preparations.

Calcium regulation of potassium fluxes in rabbit aorta during activation by noradrenaline or high potassium medium.

The dependency of noradrenaline- and depolarization-mediated increases in K permeability on cellular Ca was investigated by measuring the effect of diltiazem and Ca-free medium on stimulated 42K efflux and contracture. The increase in the rate constant (k) for 42K efflux induced by cellular depolarization with 80 mM-K was inhibited by 70% in the presence of 10(-5) M-diltiazem. The noradrenaline (NA)-mediated increase in k was only slightly suppressed by diltiazem at 10(-5) M-NA, but was inhibited by diltiazem in a dose-dependent fashion for a submaximal concentration of NA 10(-7) M. Similar inhibitory effects were observed on contractile responses. Basal 42K efflux progressively increased in a 0 Ca physiological salt solution (PSS) containing 2 mM-EGTA. This process was suppressed in a concentration-dependent manner as [Mg] was increased. In experiments utilizing 0 Ca PSS, [Mg] was therefore raised to 15 mM to maintain stable basal effluxes. Ca removal for 30 min reduced the 80 mM-K-mediated increase in k by 64%. The NA-induced increase in k became more transient in 0 Ca PSS with a progressive diminution in the magnitude of this response as the duration of Ca depletion was increased. The diminution of the 42K efflux response in 0 Ca PSS was well fitted by a mono-exponential function (half-time, t1/2 = 46 min). The NA-induced contracture in 0 Ca solution decreased with a biphasic time course subsequent to Ca removal. Intracellular Ca release by NA, measured by means of a 45Ca efflux protocol, decreased as a mono-exponential function of time, with a t 1/2 of 21 min. We conclude that both alpha-receptor activation and membrane depolarization increase K permeability largely as a result of increased cellular free [Ca]. The effect of depolarization appears to depend mainly on influx of extracellular Ca. NA increases both influx and intracellular release of Ca which serve to open K channels. The apparent release of cellular Ca as measured by 45Ca efflux into Ca-free solution decreased more rapidly, however, than did the NA stimulation of 42K efflux and tension. These observations may result from the presence of a slowly depleted Ca store which cannot be detected directly by measuring NA-induced release of 45Ca.