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1.
Ammonium Uptake by Rice Roots (II. Kinetics of 13NH4+ Influx across the Plasmalemma) 总被引:9,自引:5,他引:4
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Short-term influxes of 13NH4+ were measured in intact roots of 3-week-old rice (Oryza sativa L. cv M202) seedlings that were hydroponically grown at 2, 100, or 1000 [mu]M NH4+. Below 1 mM external concentration ([NH4+]0), influx was saturable and due to a high-affinity transport system (HATS). For the HATS, Vmax values were negatively correlated and Km values were positively correlated with NH4+ provision during growth and root [NH4+]. Between 1 and 40 mM [NH4+]0, 13NH4+ influx showed a linear response due to a low-affinity transport system (LATS). The 13NH4+ influxes by the HATS, and to a lesser extent the LATS, are energy-dependent processes. Selected metabolic inhibitors reduced influx of the HATS by 50 to 80%, but of the LATS by only 31 to 51%. Estimated values for Q10 (the ratio of rates at temperatures differing by 10[deg]C) for HATS were greater than 2.4 at root temperatures from 5 to 10[deg]C and were constant at approximately 1.5 between 5 and 30[deg]C for the LATS. Influx of 13NH4+ by the HATS was insensitive to external pH in the range from 4.5 to 9.0, but influx by the LATS declined significantly beyond pH 6.0. The data presented are discussed in the context of the kinetics, energy dependence, and the regulation of ammonium influx. 相似文献
2.
Influxes of 13NH4+ across the root plasmalemma were measured in intact seedlings of Picea glauca (Moench) Voss. Two kinetically distinct uptake systems for NH4+ were identified. In N-deprived plants, a Michaelis-Menten-type high-affinity transport system (HATS) operated in a 2.5 to 350 [mu]M range of external NH4+ concentration ([NH4 +]o). The Vmax of this HATS was 1.9 to 2.4 [mu]mol g-1 h-1, and the Km was 20 to40 [mu]M. At [NH4+]o from 500 [mu]M to 50 mM, a linear low-affinity system (LATS) was apparent. Both HATS and LATS were constitutive. A time-dependence study of NH4+ influx in previously N-deprived seedlings revealed a small transient increase of NH4+ influx after 24 h of exposure to 100 [mu]M [NH4+]o. This was followed by a decline of influx to a steady-state value after 4 d. In seedlings exposed to 100 [mu]M external NO3- concentration for 3 d, the Vmax for NH4+ uptake by HATS was increased approximately 30% compared to that found in N-deprived seedlings, whereas LATS was down-regulated. The present study defines the much higher uptake capacity for NH4+ than for N03- in seedlings of this species. 相似文献
3.
Low-affinity potassium uptake system in the archaeon Methanobacterium thermoautotrophicum: overproduction of a 31-kilodalton membrane protein during growth on low-potassium medium.
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During growth on low-K+ medium (1 mM K+), Methanobacterium thermoautotrophicum accumulated K+ up to concentration gradients ([K+]intracellular/[K+]extracellular) of 25,000- to 50,000-fold. At these gradients ([K+]extracellular of < 20 microM), growth ceased but could be reinitiated by the addition of K+ or Rb+. During K+ starvation, the levels of a protein with an apparent molecular weight of 31,000 increased about sixfold. The protein was associated with the membrane and could be extracted by detergents. Cell suspensions of M. thermoautotrophicum obtained after K+-limited growth catalyzed the transport of both K+ and Rb+ with apparent Km and Vmax values of 0.13 mM and 140 nmol/min/mg, respectively, for K+ and 3.4 mM and 140 nmol/min/mg, respectively, for Rb+. Rb+ competitively inhibited K+ uptake with an inhibitor constant of about 10 mM. Membranes of K+-starved cells did not exhibit K+-stimulated ATPase activity. Immunoblotting with antisera against Escherichia coli Kdp-ATPase did not reveal any specific cross-reactivity against membrane proteins of K+-starved cells. Cells of M. thermoautotrophicum grown at a high potassium concentration (50 mM) catalyzed K+ and Rb+ transport at similar apparent Km values (0.13 mM for K+ and 3.3 mM for Rb+) but at significantly lower apparent Vmax values (about 60 nmol/min/mg for both K+ and Rb+) compared with K+-starved cells. From these data, it is concluded that the archaeon M. thermoautotrophicum contains a low-affinity K+ uptake system which is overproduced during growth on low-K+ medium. 相似文献
4.
The mechanism of insulin stimulation of (Na+,K+)-ATPase transport activity in muscle 总被引:8,自引:0,他引:8
Since the mechanism underlying the insulin stimulation of (Na+,K+)-ATPase transport activity observed in multiple tissues has remained undetermined, we have examined (Na+,K+)-ATPase transport activity (ouabain-sensitive 86Rb+ uptake) and Na+/H+ exchange transport (amiloride-sensitive 22Na+ influx) in differentiated BC3H-1 cultured myocytes as a model of insulin action in muscle. The active uptake of 86Rb+ was sensitive to physiological insulin concentrations (1 nM), yielding a maximum increase of 60% without any change in 86Rb+ permeability. In order to determine the mechanism of insulin stimulation of (Na+,K+)-ATPase activity, we demonstrated that insulin also stimulates passive 22Na+ influx by Na+/H+ exchange transport (maximal 200% increase) and an 80% increase in intracellular Na+ concentration with an identical time course and dose-response curve as insulin-stimulated (Na+,K+)-ATPase transport activity. Incubation of the cells with high [Na+] (195 mM) significantly potentiated insulin stimulation of ouabain-inhibitable 86Rb+ uptake. The ionophore monensin, which also promotes passive Na+ entry into BC3H-1 cells, mimics the insulin stimulation of ouabain-inhibitable 86Rb+ uptake. In contrast, incubation with amiloride or low [Na+] (10 mM), both of which inhibit Na+/H+ exchange transport, abolished the insulin stimulation of (Na+,K+)-ATPase transport activity. Furthermore, each of these insulin-stimulated transport activities displayed a similar sensitivity to amiloride. These results indicate that insulin stimulates a large increase in Na+/H+ exchange transport and that the resulting Na+ influx increases the intracellular Na+ concentration, thus activating the internal Na+ transport sites of the (Na+,K+)-ATPase. This Na+ influx is, therefore, the mediator of the insulin-induced stimulation of membrane (Na+,K+)-ATPase transport activity classically observed in muscle. 相似文献
5.
Futile plasma membrane cycling of ammonium (NH4+) is characteristic of low-affinity NH4+ transport, and has been proposed to be a critical factor in NH4+ toxicity. Using unidirectional flux analysis with the positron-emitting tracer 13N in intact seedlings of barley (Hordeum vulgare L.), it is shown that rapid, futile NH4+ cycling is alleviated by elevated K+ supply, and that low-affinity NH4+ transport is mediated by a K+-sensitive component, and by a second component that is independent of K+. At low external [K+] (0.1 mM), NH4+ influx (at an external [NH4+] of 10 mM) of 92 micromol g(-1) h(-1) was observed, with an efflux:influx ratio of 0.75, indicative of rapid, futile NH4+ cycling. Elevating K+ supply into the low-affinity K+ transport range (1.5-40 mM) reduced both influx and efflux of NH4+ by as much as 75%, and substantially reduced the efflux:influx ratio. The reduction of NH4+ fluxes was achieved rapidly upon exposure to elevated K+, within 1 min for influx and within 5 min for efflux. The channel inhibitor La3+ decreased high-capacity NH4+ influx only at low K+ concentrations, suggesting that the K+-sensitive component of NH4+ influx may be mediated by non-selective cation channels. Using respiratory measurements and current models of ion flux energetics, the energy cost of concomitant NH4+ and K+ transport at the root plasma membrane, and its consequences for plant growth are discussed. The study presents the first demonstration of the parallel operation of K+-sensitive and -insensitive NH4+ flux mechanisms in plants. 相似文献
6.
We have investigated both the kinetics and regulation of 15NH4+ influx in roots of 3-month-old hydroponically grown Citrus (Citrus sinensis L. Osbeck x Poncirus trifoliata Blanco) seedlings. The 15NH4+ influx is saturable below an external ammonium concentration of 1 mM, indicating the action of a high-affinity transport system (HATS). The HATS is under feedback repression by the N status of the plant, being down-regulated in plants adequately supplied with N during growth, and up-regulated by N-starvation. When assayed between 1 and 50 mM [15NH4+]0, the 15NH4+ influx showed a linear response typical of a low-affinity transport system (LATS). The activity of the LATS increased in plants supplied with NH4+ as compared with plants grown on an N-free medium. Transfer of the plants to N-free solution resulted in a marked decrease in the LATS-mediated 15NH4+ influx. Accordingly, resupply of NH4+ after N-starvation triggered a dramatic stimulation of the activity of the LATS. These data provide evidence that in Citrus plants, the LATS or at least one of its components is inducible by NH4+. Even when up-regulated, both the HATS and the LATS displayed a limited capacity, as compared with that usually found in herbaceous species. The use of various metabolic uncouplers or inhibitors indicated that 15NH4+ influx mediated by the HATS is strongly dependent on energy metabolism and H+ transmembrane electrochemical gradient. By contrast, the LATS is not affected by protonophores or inhibitors of the H(+)-ATPase, suggesting that its activity is mostly driven by the NH4+/NH3 transmembrane gradient. In agreement with these hypotheses, the HATS-mediated 15NH4+ influx was strongly inhibited when the solution pH was raised from 4 to 7, whereas influx mediated by the LATS was slightly stimulated. 相似文献
7.
Formyl peptide stimulation of superoxide anion release from lung macrophages: sodium and potassium involvement 总被引:1,自引:0,他引:1
We examined the role of the monovalent cations Na+ and K+ in the events encompassing the release of O-2 by alveolar macrophages after stimulation with formyl methionyl phenylalanine (FMP). This was accomplished by determining the effect of changing the extracellular [Na+] and/or [K+] on FMP-stimulated O-2 production; and measuring 22Na+, 42K+ and 86Rb+ influx and efflux and intracellular [K+] for control and FMP-stimulated alveolar macrophages. Stimulated O-2 production was relatively insensitive to changes in extracellular K+ or Na+ concentrations until the [Na+] was decreased below 35 mM. At 4 mM [Na+], the rate of O-2 production remained at 75% of the maximal rate observed at physiological concentrations of [Na+]. Both influx and efflux of 22Na+ were stimulated above control rates by FMP. The increased rates of fluxes lasted for a few minutes suggesting a transient increase in membrane permeability to Na+. Ouabain partially inhibited 22Na+ efflux but had no effect on O-2 release. The influx of 86Rb+ and 42K+ was not altered by the addition of FMP but was virtually abolished in the presence of 10 microM ouabain or 1 mM quinine. In the presence of extracellular calcium, FMP-stimulated a prolonged (greater than 20 minutes) increase in 86Rb+ or 42K+ efflux which was inhibitable by 1 mM quinine. In the absence of extracellular calcium, FMP stimulation of K+ efflux was greatly diminished and was not affected by quinine, although quinine still inhibited O-2 production under these conditions. It was also observed that there was a loss of intracellular K+ when cells were stimulated by FMP in the presence of Ca+2, but not in the absence of Ca+2. Taken together, these results suggest a minimal direct role, if any, for K+ in the events that lead to FMP-stimulated O-2 release by alveolar macrophages. 相似文献
8.
Ouabain at nanomolar concentrations stimulates total Rb+ influx by 20 +/- 2% in monolayer cultures of myocytes which were either in physiologic ionic steady-state conditions ('control') or 'loaded with Na+' following exposure to K+-free medium. The ouabain-stimulated Rb+ influx was completely abolished by 0.1 mM bumetanide both in 'control' and in 'Na+-loaded' myocytes. Thus, addition of nanomolar concentrations of ouabain to myocytes markedly stimulate the bumetanide-sensitive Rb+ influx. This influx was increased up to 3- and 4-fold in 'control' and 'Na+-loaded' myocytes, respectively. Ouabain at nanomolar concentrations had no significant effect on the component of 86Rb+ influx which is inhibited by millimolar concentrations of ouabain (the so called 'ouabain-sensitive' or 'pump-mediated' Rb+ influx) in 'control' and 'Na+-loaded' cells. It is proposed that the increased rates of bumetanide-sensitive Rb+ influx are accompanied by an increased bumetanide-sensitive Na+ influx through the Na+/K+ cotransporter and thus to a transient increase in intracellular Na+ concentrations [Na+]i. The increase in [Na+]i, subsequently causes a transient elevation in [Ca2+]i via the Na+/Ca2+ exchanger and may be involved in the regulation of cardiac cells' contractility. 相似文献
9.
42K+ and 86Rb+ were used to determine the subcellular distribution of potassium in Chlamydomonas reinhardtii by compartmental analysis. In both wild type and a mutant strain, three distinct compartments (referred to as I, II, and III) were apparent. Using 42K+, we found that these had half-lives for K+ exchange of 1.07 min, 12.8 min, and 2.9 h, respectively, in wild-type cells and 0.93 min, 14.7 min, and 9.8 h, respectively, for the mutants. Half-lives were not significantly different when 86Rb+ was used to trace K+. Compartments I and II probably correspond to the cell wall and cytoplasm, respectively. Based on the lack of a large central vacuole in Chlamydomonas, the effect of a dark pretreatment on the kinetic properties of compartment III and the similarity between the [K+] of compartment III and that of isolated chloroplasts, this slowly exchanging compartment was identified as the chloroplast. Growth of wild-type cells at 100 [mu]M (instead of 10 mM K+) caused no change of cytoplasmic [K+] but reduced chloroplast [K+] very substantially. The mutants failed to grow at 100 [mu]M K+. 相似文献
10.
Agonist-specific regulation of [Na+]i in pancreatic acinar cells 总被引:1,自引:1,他引:0
《The Journal of general physiology》1995,106(6):1243-1263
In a companion paper (Zhao, H., and S. Muallem. 1995), we describe the relationship between the major Na+,K+, and Cl- transporters in resting pancreatic acinar cells. The present study evaluated the role of the different transporters in regulating [Na+]i and electrolyte secretion during agonist stimulation. Cell stimulation increased [Na+]i and 86Rb influx in an agonist-specific manner. Ca(2+)-mobilizing agonists, such as carbachol and cholecystokinin, activated Na+ influx by a tetraethylammonium-sensitive channel and the Na+/H+ exchanger to rapidly increase [Na+]i from approximately 11.7 mM to between 34 and 39 mM. As a consequence, the NaK2Cl cotransporter was largely inhibited and the activity of the Na+ pump increased to mediate most of the 86Rb(K+) uptake into the cells. Secretin, which increases cAMP, activated the NaK2Cl cotransporter and the Na+/H+ exchanger to slowly increase [Na+]i from approximately 11.7 mM to an average of 24.6 mM. Accordingly, secretin increased total 86Rb uptake more than the Ca(2+)- mobilizing agonists and the apparent coupling between the NaK2Cl cotransport and the Na+ pump. All the effects of secretin could be attributed to an increase in cAMP, since forskolin affected [Na+]i and 86Rb fluxes similar to secretin. The signaling pathways mediating the effects of the Ca(2+)-mobilizing agonists were less clear. Although an increase in [Ca2+]i was required, it was not sufficient to account for the effect of the agonists. Activation of protein kinase C stimulated the NaK2Cl cotransporter to increase [Na+]i and 86Rb fluxes without preventing the inhibition of the cotransporter by Ca(2+)-mobilizing agonists. The effects of the agonists were not mediated by changes in cell volume, since cell swelling and shrinkage did not reproduce the effect of the agonists on [Na+]i and 86Rb fluxes. The overall findings of the relationships between the various Na+,K+, and Cl- transporters in resting and stimulated pancreatic acinar cells are discussed in terms of possible models of fluid and electrolyte secretion by these cells. 相似文献
11.
Triads and transverse tubules isolated from mammalian skeletal muscle actively accumulated Na+ in the presence of K+ and Mg-ATP. Active Na+ transport exhibited a fast single-exponential phase, lasting 2 min, followed by slower linear uptake that continued for 10 minutes. Valinomycin stimulated Na+ uptake, suggesting it decreased a pump-generated membrane potential gradient (Vm) that prevented further Na+ accumulation. At the end of the fast uptake phase transverse tubule vesicles incubated in 30 mM external [Na+] attained a ratio [Na+]in/[Na+]out=13.4. From this ratio and the transverse tubule volume of 0.35 microl/mg protein measured in this work, [Na+]in=400 mM was calculated. Determinations of active K+ transport in triads, using 86Rb+ as tracer, showed a 30% decrease in vesicular 86Rb+ content two minutes after initiating the reaction, followed by a slower uptake phase during which vesicles regained their initial 86Rb+ content after 10 minutes. Transverse tubule volume increase during active Na+ transport-as shown by light scattering changes of isolated vesicles--presumably accounted for the secondary Na+ and 86Rb+ uptake phases. These combined results indicate that isolated triads have highly sealed transverse tubules that can be polarized effectively by the Na+ pump through the generation of significant Na+ gradients. 相似文献
12.
Rapid, futile K+ cycling and pool-size dynamics define low-affinity potassium transport in barley 总被引:7,自引:0,他引:7
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Using the short-lived radiotracer 42K+, we present a comprehensive subcellular flux analysis of low-affinity K+ transport in plants. We overturn the paradigm of cytosolic K+ pool-size homeostasis and demonstrate that low-affinity K+ transport is characterized by futile cycling of K+ at the plasma membrane. Using two methods of compartmental analysis in intact seedlings of barley (Hordeum vulgare L. cv Klondike), we present data for steady-state unidirectional influx, efflux, net flux, cytosolic pool size, and exchange kinetics, and show that, with increasing external [K+] ([K+]ext), both influx and efflux increase dramatically, and that the ratio of efflux to influx exceeds 70% at [K+]ext > or = 20 mm. Increasing [K+]ext, furthermore, leads to a shortening of the half-time for cytosolic K+ exchange, to values 2 to 3 times lower than are characteristic of high-affinity transport. Cytosolic K+ concentrations are shown to vary between 40 and 200 mm, depending on [K+]ext, on nitrogen treatment (NO3- or NH4+), and on the dominant mode of transport (high- or low-affinity transport), illustrating the dynamic nature of the cytosolic K+ pool, rather than its homeostatic maintenance. Based on measurements of trans-plasma membrane electrical potential, estimates of cytosolic K+ pool size, and the magnitude of unidirectional K+ fluxes, we describe efflux as the most energetically demanding of the cellular K+ fluxes that constitute low-affinity transport. 相似文献
13.
Regulatory interaction of ATP Na+ and Cl- in the turnover cycle of the NaK2Cl cotransporter 总被引:2,自引:2,他引:0
《The Journal of general physiology》1993,101(6):889-908
To probe the mechanism by which intracellular ATP, Na+, and Cl- influence the activity of the NaK2Cl cotransporter, we measured bumetanide-sensitive (BS) 86Rb fluxes in the osteosarcoma cell line UMR- 106-01. Under physiological gradients of Na+, K+, and Cl-, depleting cellular ATP by incubation with deoxyglucose and antimycin A (DOG/AA) for 20 min at 37 degrees C reduced BS 86Rb uptake from 6 to 1 nmol/mg protein per min. Similar incubation with 0.5 mM ouabain to inhibit the Na+ pump had no effect on the uptake, excluding the possibility that DOG/AA inhibited the uptake by modifying the cellular Na+ and K+ gradients. Loading the cells with Na+ and depleting them of K+ by a 2-3- h incubation with ouabain or DOG/AA increased the rate of BS 86Rb uptake to approximately 12 nmol/mg protein per min. The unidirectional BS 86Rb influx into control cells was approximately 10 times faster than the unidirectional BS 86Rb efflux. On the other hand, at steady state the unidirectional BS 86Rb influx and efflux in ouabain-treated cells were similar, suggesting that most of the BS 86Rb uptake into the ouabain-treated cells is due to K+/K+ exchange. The entire BS 86Rb uptake into ouabain-treated cells was insensitive to depletion of cellular ATP. However, the influx could be converted to ATP-sensitive influx by reducing cellular Cl- and/or Na+ in ouabain-treated cells to impose conditions for net uptake of the ions. The BS 86Rb uptake in ouabain-treated cells required the presence of Na+, K+, and Cl- in the extracellular medium. Thus, loading the cells with Na+ induced rapid 86Rb (K+) influx and efflux which, unlike net uptake, were insensitive to cellular ATP. Therefore, we suggest that ATP regulates a step in the turnover cycle of the cotransporter that is required for net but not K+/K+ exchange fluxes. Depleting control cells of Cl- increased BS 86Rb uptake from medium-containing physiological Na+ and K+ concentrations from 6 to approximately 15 nmol/mg protein per min. The uptake was blocked by depletion of cellular ATP with DOG/AA and required the presence of all three ions in the external medium. Thus, intracellular Cl- appears to influence net uptake by the cotransporter. Depletion of intracellular Na+ was as effective as depletion of Cl- in stimulating BS 86Rb uptake.(ABSTRACT TRUNCATED AT 400 WORDS) 相似文献
14.
Yu M Lopez B Dos Santos EA Falck JR Roman RJ 《American journal of physiology. Regulatory, integrative and comparative physiology》2007,292(6):R2400-R2405
Previous studies have indicated that 20-hydroxyeicosatetraenoic acid (20-HETE) inhibits Na+ transport in the medullary thick ascending loop of Henle (mTALH), but the mechanisms involved remain uncertain. The present study compared the effects of 20-HETE with those of ouabain and furosemide on intracellular Na+ concentration ([Na+]i), Na+ -K+ -ATPase activity, and 86Rb+ uptake, an index of Na+ transport, in mTALH isolated from rats. Ouabain (2 mM) increased, whereas furosemide (100 microM) decreased, [Na+]i in the mTALH of rats. Ouabain and furosemide inhibited 86Rb+ uptake by 91 and 30%, respectively. 20-HETE (1 microM) had a similar effect as ouabain and increased [Na+]i from 19 +/- 1 to 30 +/- 1 mM. 20-HETE reduced Na+ -K+ -ATPase activity by 30% and 86Rb+ uptake by 37%, but it had no effect on 86Rb+ uptake or [Na+]i in the mTALH of rats pretreated with ouabain. 20-HETE inhibited 86Rb+ uptake by 12% and increased [Na+]i by 19 mM in mTALH pretreated with furosemide. These findings indicate that 20-HETE secondarily inhibits Na+ transport in the mTALH of the rat, at least, in part by inhibiting the Na+ -K+ -ATPase activity and raising [Na+]i. 相似文献
15.
Na+, K+, and Cl- transport in resting pancreatic acinar cells 总被引:2,自引:1,他引:1
《The Journal of general physiology》1995,106(6):1225-1242
To understand the role of Na+, K+, and Cl- transporters in fluid and electrolyte secretion by pancreatic acinar cells, we studied the relationship between them in resting and stimulated cells. Measurements of [Cl-]i in resting cells showed that in HCO3(-)-buffered medium [Cl- ]i and Cl- fluxes are dominated by the Cl-/HCO3- exchanger. In the absence of HCO3-, [Cl-]i is regulated by NaCl and NaK2Cl cotransport systems. Measurements of [Na+]i showed that the Na(+)-coupled Cl- transporters contributed to the regulation of [Na+]i, but the major Na+ influx pathway in resting pancreatic acinar cells is the Na+/H+ exchanger. 86Rb influx measurements revealed that > 95% of K+ influx is mediated by the Na+ pump and the NaK2Cl cotransporter. In resting cells, the two transporters appear to be coupled through [K+]i in that inhibition of either transporter had small effect on 86Rb uptake, but inhibition of both transporters largely prevented 86Rb uptake. Another form of coupling occurs between the Na+ influx transporters and the Na+ pump. Thus, inhibition of NaK2Cl cotransport increased Na+ influx by the Na+/H+ exchanger to fuel the Na+ pump. Similarly, inhibition of Na+/H+ exchange increased the activity of the NaK2Cl cotransporter. The combined measurements of [Na+]i and 86Rb influx indicate that the Na+/H+ exchanger contributes twice more than the NaK2Cl cotransporter and three times more than the NaCl cotransporter and a tetraethylammonium-sensitive channel to Na+ influx in resting cells. These findings were used to develop a model for the relationship between the transporters in resting pancreatic acinar cells. 相似文献
16.
On incubation of HeLa cells in chilled isotonic medium, intracellular Na+ (Nac+) increased and K+ (Kc+) decreased with time, reaching steady levels after 3 h. The steady levels varied in parallel with the extracellular cation concentrations ([Na+]e, [K+]e). The cell volumes and the protein and water contents, respectively, of cells kept for 3 h in chilled media of various [Na+]e and [K+]e were not significantly different. Ouabain-sensitive Rb+ influx took place at the initial rate for a certain period which depended on [Na+]c at the beginning of the assays. The existence of two external K+ loading sites per Na+/K+-pump was demonstrated. The affinities of the sites for Rb+ as a congener of K+ were almost the same. Na+e inhibited ouabain-sensitive Rb+ influx competitively, whereas K+ was not inhibitory. Kinetic parameters were determined: the K 1/2 for Rbe+ in the absence of Na+e was 0.16 mM and th Ki for Na+e was 36.8 mM; the K 1/2 for Na+c was 19.5 mM and the Ki for K+c seemed to be extremely large. The rate equation of the ouabain-sensitive Rb+ influx suggests that Na+ and K+ are exchanged alternately through the pump by a binary mechanism. 相似文献
17.
Microdissected, beta-cell-rich pancreatic islets from ob/ob mice were used in studies of 86Rb+ transport. D-Glucose (20 mM) induced a biphasic reduction in 86Rb+ efflux. The reduction stabilized within 10 min at 34% of the efflux rate at zero glucose. The initial 86Rb+ uptake (5 min) was dose-dependently reduced by ouabain with maximum inhibition at 1 mM. D-Glucose (20 mM) did not affect the ouabain-sensitive 86Rb+ influx but markedly reduced (48%) the ouabain-resistant isotope influx. The results suggest that D-glucose does not affect the Na+/K+ pump in pancreatic beta-cells and that the glucose-sensitive K+-transporting modalities (K+ channels) in the beta-cells can mediate both inward and outward K+ flux. 相似文献
18.
M C Montero J Bolufer A Ilundáin 《Comparative biochemistry and physiology. A, Comparative physiology》1991,99(1-2):31-36
1. Efflux of K+ was measured in pre-loaded (86Rb+) chicken enterocytes incubated in buffers with external K+ concentration ([K+]0) between 1 and 40 mM. 2. A decrease in [K+]0 from 6 to 1 mM reduced the rate constant of K+ efflux, whereas it was stimulated by increasing [K+]0 from 6 to 40 mM. 3. The inhibitory effect of low [K+]0 on K+ efflux was: (i) higher than that expected from a change in the electrical driving force, suggesting that membrane K+ permeability has been decreased, and (ii) attenuated by A23187 and Na(+)-free buffers. 4. The effect of A23187 on K(+)-induced K+ efflux was abolished by apamin and that of Na(+)-free buffers by apamin, quinine or verapamil, which suggests that the effect of low K+ on K+ efflux seems to be due to decreased intracellular Ca2+ concentration. 5. The stimulatory effect of 40 mM K0+ on K+ exit can be accounted for by an increase in the electrical driving force. 6. The efflux of K+ at 40 mM K0 appears to occur through Ca2(+)-activated K+ channels (KCa) since it was prevented by 500 microM quinine and unaffected by bumetanide or 3,4-diaminopyridine. 7. In addition, the current results show that an increase in external K+ concentration reduced the ability of quinine to inhibit KCa channels, and even abolished that of Ba2+ and apamin. 相似文献
19.
Discrimination between Rb+ and K+ by Escherichia coli. 总被引:1,自引:0,他引:1
1. The K+ requirment of Escherichia coli is only partially fulfilled by Rb+. The molar growth yield on Rb+ was about 5% of that on K+ and the growth rate in Rb+-supplemented media is lower thatn in K+ influx by any of the four K+ transport systems of E. coli. The high-affinity Kdp system (Km = 2 micron) is poorly traced by 86Rb+. It discriminates against a 86Rb+ tracer at least 1000-fold. The two moderate affinity systems, the high-rate TrkA system (Km = 1.5 mM) and the moderate rate TrkD system (Km = 0.5 mM), discriminate against a 86Rb+ tracer by approximately 10-fold and 25-fold, respectively. 86Rb+ is preferred by the low-rate TrkF system and overestimates its K+ influx by 40%. 相似文献
20.
The modulation of rat brain Na(+)-Ca2+ exchange by K+ 总被引:1,自引:0,他引:1
The involvement of potassium ions in the Na(+)-Ca2+ exchange process was studied in rat brain synaptic plasma membrane (SPM) vesicles. Addition of equimolar [K+] to the intravesicular and the extravesicular medium led to a stimulation of the Na+ gradient-dependent Ca2+ influx; this stimulation was noticeable already at 0.5 mM and reached its maximum at 2 mM K+. The magnitude of the K+ stimulation was between 1.3-2.5-fold in different SPM preparations. K+ ions also stimulated the Na(+)-dependent Ca2+ efflux. K+ stimulation of Na(+)-Ca2+ exchange is of considerable specificity, since it is not mimicked by either Li+ or H+. The following lines of evidence suggest that K+ modulation of Na(+)-Ca2+ exchange involves the catalytic moiety of the transporter itself and not an unrelated K+ channel which modulates the membrane potential. 1) K+ stimulation of the transport process was conserved following reconstitution of the transporter into phospholipid-rich liposomes, an experimental condition which presumably separates the native membrane proteins among different vesicular structures. 2) K+ stimulation of Na+ gradient-dependent Ca2+ influx persists also when the build up of negative inside membrane potential is prevented by addition of carbonyl cyanide p-trifluoromethoxy phenylhydrazone which renders the membrane highly permeable to protons both in the native and the reconstituted preparation. 3) K+ stimulation of Na+ gradient-dependent Ca2+ influx is obtained also when tetraethylammonium chloride, 2,3-diaminopyridine and Cs+ are added to the Ca2+ uptake medium. Reconstituted SPM vesicles take up 86Rb+ in response to activation of Na+ gradient-dependent Ca2+ influx. The ratio of Ca2+ taken up by SPM vesicles in a Na+ gradient-dependent manner to the corresponding amounts of Rb+ taken up varies between 8-5 in different SPM preparations. If the stoichiometry of the process is 1 Rb+/1 Ca2+, then Rb+ cotransport is mediated by 10-20% of the transporters present in the preparation. 相似文献