Na+, K+ and Cl- in Xylem Sap Flowing to Shoots of NaCl-Treated Barley

Munns, R. 1985. Na⁺, K⁺ and Cl^– in xylem sap flowing toshoots of NaCl-treated barley.—J. exp. Bot. 36: 1032–1042. Na⁺, Cl^– and K⁺ concentrations were measured in xylemsap obtained by applying pressure to the roots of decapitatedbarley plants grown at external [NaCl] of 0, 25, 50, 100, 150and 200 mol m^–3. For any given NaCl treatment, ion concentrationsin the xylem sap were hyperbolically related to the flux ofwater. Ion concentrations in sap collected at very low volumefluxes (without applied pressure) were 5–10 times higherthan in sap collected at moderate fluxes (under pressure). Fora given moderate volume flux, Na+ concentration in the xylemsap, [Na⁺]_x, was only 4.0 mol m^–3 at external [NaCl] of25–150 mol m^–3, and increased to 7.0 mol m^–3at 200 mol m^–3. [Cl-]x showed a similar pattern. Thisshows there would be little difference in the rate of uptaketo the shoot of plants at 25–150 mol m^–3 externalNaCl and indicates little change even at 200 mol m^-3 NaCl becausetranspiration rates would be much lower. Thus the reduced growthof the shoot of plants at high NaCl concentrations is not dueto higher uptake rates of Na⁺ or Cl^–. The fluxes of Na⁺, Cl^– and K^– increased non-linearlywith increasing volume flux indicating little movement of saltin the apoplast. The flux of K⁺ increased even when [K⁺]_x wasgreater than external [K⁺], indicating that membrane transportprocesses modify the K⁺ concentration in the transpiration streamas it flows through the root system. Key words: -Xylem sap, Na⁺, K⁺, Cl^– fluxes, salinity, barley     

Cl- secretory effects of EBIO in the rabbit conjunctival epithelium

Experiments were conducted to determine whether the Cl^– secretagogue, 1-ethyl-2-benzimidazolinone (EBIO), stimulates Cl^– transport in the rabbit conjunctival epithelium. For this study, epithelia were isolated in an Ussing-type chamber under short-circuit conditions. The effects of EBIO on the short-circuit current (I_sc) and transepithelial resistance (R_t) were measured under physiological conditions, as well as in experiments with altered electrolyte concentrations. Addition of 0.5 mM EBIO to the apical bath stimulated the control I_sc by 64% and reduced R_t by 21% (P < 0.05; paired data). Under Cl^–-free conditions, I_sc stimulation using EBIO was markedly attenuated. In the presence of an apical-to-basolateral K⁺ gradient and permeabilization of the apical membrane, the majority of the I_sc reflected the transcellular movement of K⁺ via basolateral K⁺ channels. Under these conditions, EBIO in combination with A23187 elicited nearly instantaneous 60–90% increases in I_sc that were sensitive to the calmodulin antagonist calmidazolium and the K⁺ channel blocker tetraethyl ammonium. In the presence of an apical-to-basolateral Cl^– gradient and nystatin permeabilization of the basolateral aspect, EBIO increased the Cl^–-dependent I_sc, an effect prevented by the channel blocker glibenclamide (0.3 mM). The latter compound also was used to determine the proportion of EBIO-evoked unidirectional ³⁶Cl^– fluxes in the presence of the Cl^– gradient that traversed the epithelium transcellularly. Overall, EBIO activated apical Cl^– channels and basolateral K⁺ channels (presumably those that are Ca²⁺ dependent), thereby suggesting that this compound, or related derivatives, may be suitable as topical agents to stimulate fluid transport across the tissue in individuals with lacrimal gland deficiencies. Ussing chamber; short-circuit current; electrolyte transport; chloride secretagogue; potassium conductance; 1-ethyl-2-benzimidazolinone; 1,10-phenanthroline     

Characterization of the Ca2+-Dependent Cl- Efflux in Perfused Chara Cells

The mechanism of the Ca²⁺-dependent Cl^– efflux was studiedin tonoplast-free cells, in which the intracellular chemicalcomposition can be freely controlled. Tonoplast-free cells wereprepared by perfusing the cell interior of internodal cellsof Chara corallina with a medium that contained EGTA. The Ca²⁺-inducedCl^– efflux was measured together with the membrane potentialduring continuous intracellular perfusion. The dependenciesof Cl^– efflux and the membrane potential on the intracellularCa²⁺ or Cl^– concentrations were analyzed. When perfusionwas started with medium that contained Ca²⁺ ions, Cl^–efflux and membrane depolarization were induced. The amountof Cl^– efflux varied considerably among individual cells.The rate of efflux decreased exponentially but a residual effluxremained detectable. The Cl^– efflux was induced at concentrationsof Ca²⁺ ions above 1 µM and reached a maximum at 1 mM.By contrast, the membrane depolarization reached a maximum atabout 10 µM Ca²⁺. The rate of Cl^– efflux increasedlinearly with logarithmic increases in the intracellular Cl^–concentrations. These findings suggest that more than two kindsof Ca²⁺-dependent Cl^– channel might be present in theplasma membrane. Addition of ATP or its removal from the perfusion medium didnot affect the Ca²⁺-dependent Cl^– efflux. Calmodulin antagonistsslightly inhibited the Ca²⁺-dependent Cl^– efflux. ¹Present address: Biological Laboratory, Hitotsubashi University,Naka 2-1, Kunitachi, Tokyo, 186 Japan.     

Chloride Transport in Chara corallina and the Electrochemical Potential Difference for Hydrogen Ions

The pH of the cytoplasm of Chara corallina cells has been measuredwith the weak acid 5,5-dimethyloxazolidine-2,4-dione (DM0).Over an external pH range 4·5–9·5 the resultsfit the regression equation pH_cytoplasm=6·28+0·22pH_out. Using measured values of the electric potential difference acrossthe plasmalemma we have calculated the electrochemical potentialdifference across this membrane for H⁺ and Cl^–. Thesedata are used to test the hypothesis that the inward transportof Cl^– is coupled to the inthix of H⁺ or, which comesto the same thing, efflux of OH^–. One-for-one couplingwill not give net Cl^– uptake from solutions with pH greaterthan about 7·2, unless the cytoplasmic Cl^– concentrationis lower than 10 mM, or the pH just outside the membrane islower than that in the bulk solution. It is shown that net Cl^–uptake proceeds from solutions with pH up to 9. The alternative possibility is that Cl^– transport is broughtabout by co-transport of two H⁺ for each Cl^–; this isnot ruled out by the results reported. Such a mechanism mightbe detectable by its electrogenic effect: although such effectshave not been detected, it is shown that they would be smallunder most conditions. Other possible mechanisms are discussed.     

Effects of Ammonia and Methylamine on Cl- Transport and on the pH Changes and Circulating Electric Currents Associated with HCO3- Assimilation in Chara corallina

Low concentrations of ammonia and methylamine greatly increaseCl^– influx into Chara corallina. Both amines have theirmaximum effect at pH 6.5–7.5. The amine stimulation ofCl^– influx is small below about pH 5.5. Above pH 8.5 theremay be inhibition of influx by amines. Concentrations of 10–25µM ammonia are sufficient to cause the maximum stimulationof Cl^– influx; the corresponding methylamine concentrationsare 0.1–0.2 mM. It is concluded that entry of amine cations(NH₄^$ and CH₃NH₃^$), rather than unionized bases (NH₃ and CH₃NH₂),causes Cl^– transport to be increased. Increases in rates of Cl^– transport are not necessarilyaccompanied by effects on HCO₃^$ assimilation and OH^– efflux.Measurements of localized pH differences at the cell surfaceand of circulating electric currents in the bathing solutionshow that these phenomena are only significantly affected byammonia at or above 50 µM and by methylamine at or above1.0 mM. The significance of the effects of amines is assessedin relation to current ideas about transport of Cl^–, HCO₃^–,and OH^–.     

Light-dependent potassium uptake by Pisum sativum leaf fragments

A net K⁺ influx into chopped pea leaves bathed in 5 mM KCl,0.26 M sucrose and illuminated with 4000 lux amounted to about7.5 µmoles/g fresh weight-hr, while essentially no netflux occurred in the dark. This light-dependent K⁺ uptake waslinear with time for nearly 2 hr and continuously increasedas the light intensity was raised to 9000 lux. Over half ofthe K⁺ uptake was balanced by H⁺ release into the bathing solution,possibly by a mechanism in which bicarbonate was the anion accompanyingK⁺. The replacement of Cl^– by HCO₃^– increased thelight-dependent K⁺ uptake to 56 µmoles/g fresh weight-hr.About 23% of the light-dependent K⁺ uptake in 5 mM KCl was accompaniedby a Cl^– uptake. This net Cl^– influx was less sensitiveto the uncoupler tri-Fl-CCP and more sensitive to DCMU in thebathing solution than was the K⁺ uptake. The remaining net K⁺influx into pea leaf fragments was balanced by effluxes of sodium(accounting for 5%), magnesium (8%) and calcium (1%). (Received March 31, 1969; )     

Control of Plasma Membrane Cl- Fluxes in Chara corallina by External Cl- and Light

The efflux of Cl^– at the plasma membrane of Chara wasstudied in relation to two treatments known to affect the flux:that of removal of external Cl^– and of light. It is shownthat although removal of external Cl^– results in a rapidreduction in Cl^– efflux (consistent with a direct effectof external Cl^– on the transport system) the magnitudeof this reduction in the dark is greater than the measured darkinflux. Therefore, in the dark at least, it is proposed that1:1 exchange diffusion cannot account for the trans-stimulationof efflux by external Cl^–. Light induces an inhibition of efflux and a concomitant stimulationof Cl^– influx at 20 °C, but at 10 °C the responsesto light of the two fluxes can be separated temporally. It istherefore suggested that the fluxes are not reciprocally dependenton the same factor which mediates in the light response. Furtherconsiderations show that it is unlikely that the cytoplasmicCl^– concentration mediates in the light response of eitherflux, but that changes in cytoplasmic pH may do so.     

Characteristics of the Cl- Action Site in the O2 Evolving Reaction in PS II Particles: Electrostatic Interaction with Ions

The role of Cl^– in the reactivation of O₂ evolution inphotosystem II (PS II) particles derived from spinach chloroplastswas studied in the presence of various salts. Multivalent ion(especially anion) salts were found to strongly suppress thereactivation of O₂ evolution by Cl^– in the Cl^–-depletedPS II particles in a competitive manner. The effectiveness ofanions in the suppression of Cl^–-supported O₂ evolutionwas in the order of trivalent>divalent>monovalent ones.Multivalent anions similarly suppressed O₂ evolution in theuntreated PS II particles under low and moderate Cl^– concentrations.pH dependence of the Cl^–-affinity (K_m) value for Cl^–)was also studied. Within the pH range 5.5 to 8 the K_m valuebecame higher as the pH of the medium increased. These resultssuggest that the membrane surface in the vicinity of the Cl^–action site is net positively charged and attracts Cl^–electrostatically, and that the site is almost freely accessibleto various anions. The origin and role of the local net positivedomain and the role of peripheral proteins are discussed. (Received May 27, 1985; Accepted October 8, 1985)     

Protective role of extracellular chloride in fatigue of isolated mammalian skeletal muscle

A possible role of extracellular Cl^– concentration ([Cl^–]_o) in fatigue was investigated in isolated skeletal muscles of the mouse. When [Cl^–]_o was lowered from 128 to 10 mM, peak tetanic force was unchanged, fade was exacerbated (wire stimulation electrodes), and a hump appeared during tetanic relaxation in both nonfatigued slow-twitch soleus and fast-twitch extensor digitorum longus (EDL) muscles. Low [Cl^–]_o increased the rate of fatigue 1) with prolonged, continuous tetanic stimulation in soleus, 2) with repeated intermittent tetanic stimulation in soleus or EDL, and 3) to a greater extent with repeated tetanic stimulation when wire stimulation electrodes were used rather than plate stimulation electrodes in soleus. In nonfatigued soleus muscles, application of 9 mM K⁺ with low [Cl^–]_o caused more rapid and greater tetanic force depression, along with greater depolarization, than was evident at normal [Cl^–]_o. These effects of raised [K⁺]_o and low [Cl^–]_o were synergistic. From these data, we suggest that normal [Cl^–]_o provides protection against fatigue involving high-intensity contractions in both fast- and slow-twitch mammalian muscle. This phenomenon possibly involves attenuation of the depolarization caused by stimulation- or exercise-induced run-down of the transsarcolemmal K⁺ gradient. potassium; skeletal muscle contraction; membrane potential; myotonia     

Diurnal K+ and Anion Transport in Phaseolus Pulvinus

Diurnal movement of Phaseolus leaf is caused by deformationof the laminar pulvinus located at the joint of the leaf bladeand the petiole. The plants were cultured in solutions withvarious ion compositions, and changes of K⁺, Na⁺, Ca²⁺, Mg²⁺,Cl^–, NO₃– and P₁ concentrations both in the upperand lower parts of the laminar pulvinus were measured. Culturein 10 mM KCl solution caused an increase in K⁺ and Cl^–concentrations both in the upper and lower parts without anysignificant change in the concentration of NO₃^–; culturein 10 mM KNO₃ solution caused an increase in K⁺ and NO₃^–concentration without any significant change in the concentrationof Cl^–; and culture in 10 mM KH₂PO₄ solution caused anincrease in K⁺ and P₁ concentrations without any significantchange in the concentrations of NO₃- and Cl^–. K⁺ moved from the upper to lower parts or from the lower toupper parts diurnally in all plants cultured in any solutionmentioned above. The main inorganic anion that accompanied thisK⁺ movement was Cl^– in KCl solution, and NO₃^– inKNO₃ solution. When the seedlings were cultured in distilledwater or in KH₂PO₄ solution, neither Cl^– NO₃^– norP₁ accompanied this K⁺ movement. In these cases, mainly H⁺ and/ororganic anions are supposed to move in exchange for and/or incombination with K⁺ movement. (Received November 8, 1982; Accepted June 13, 1983)     

Membrane Potential Measurement of Protoplasts Isolated from Vigna mungo Hypocotyl Using a Fluorescent Probe, diS-C3-(5)

Membrane potentials of protoplasts isolated from Vigna mungohypocotyl segments were measured using the fluorescent probediS-C₃-(5). The fluorescence intensity changed in response tothe external K⁺ concentration. Membrane potential was estimatedto be inside negative (–85?8 mV at 0.1 mM KCl) from theNernst equation for K₊. The membrane potential was not affectedby DCCD (50 µM) or low temperature (5?C). Addition of0.5 mM Ca₂₊ to the protoplast suspension markedly depolarizedthe membrane potential, and subsequent EDTA treatment repolarizedit to the initial level. The Ca₂₊ effect on the membrane potentialmay be due to change in the permeability ratio of Cl^–to K₊. (Received December 16, 1986; Accepted April 22, 1987)     

Stimulatory Effect of Chloride Ions on NADP Malic Enzyme from Leaves of two C4 Species of Cyperus

NADP malic enzyme (EC 1.1.1.40 [EC] ) from leaves of two C₄ speciesof Cyperus (C. rotundus and C. brevifolius var leiolepis) exihibiteda low level of activity in an assay mixture that contained lowconcentrations of Cl^–. This low level of activity wasmarkedly enhanced by increases in the concentration of NaClup to 200 mM. Since the activity of NADP malic enzyme was inhibitedby Na₂SO₄ and stimulated by relatively high concentration ofTris-HCl (50–100 mM, pH 7–8), the activation ofthe enzyme by NaCl appears to be due to Cl^–. Variationsin the concentration of Mg²⁺ affected the K_A (the concentrationof activator giving half-maximal activation) for Cl^–,which decreased from 500 mM to 80 mM with increasing concentrationsof Mg²⁺ from 0.5 mM to 7 mM. The K_m for Mg²⁺ was decreased from7.7 mM to 1.3 mM with increases in the concentration of NaClfrom zero to 200 mM, although the increase of V_max was not remarkable.NADP malic enzyme from Cyperus, being similar to that from otherC₄ species, was able to utilize Mn²⁺. The K_m for Mn²⁺ was 5mM, a value similar to that for Mg²⁺. The addition of 91 mMNaCl markedly decreased the K_m for Mn²⁺ to 20 +M. NADP malicenzyme from Setaria glauca, which contains rather less Cl^–than other C₄ species, was inactivated by concentrations ofNaCl above 20 mM, although slight activation of the enzyme wasobserved at low concentrations of NaCl at pH7.6. (Received February 20, 1989; Accepted June 12, 1989)     

Effects of Abscisic Acid and Cytoplasmic pH on Potassium and Chloride Efflux in Arabidopsis thaliana Seedlings

The effects of ABA, isobutyric acid (IBA) and nicotine on K⁺and Cl⁺ efflux were studied in Arabidopsis thaliana seedlings,and the role of pH_cyt, and E_m in the regulation of the effluxof these ions was discussed. The data show that treatments withIBA and nicotine influenced in opposite directions the effluxof either K⁺ or Cl^–: K⁺ efflux was increased by nicotineand reduced in the presence of IBA, whereas Cl^– effluxwas stimulated by IBA and decreased by nicotine treatment. Underall the conditions tested ABA induced cytoplasmic acidificationand inhibition of K⁺ and Cl^– net efflux. Experiments aimedto estimate the individual contribution of pH_cyt and E_m in modulatingK^– efflux indicated that, within the range of acidic pH_cytvalues, a regulation of K⁺ efflux was imposed by pH_cyt on thecontrol exerted by E_m, the efflux being inhibited by lower pH_cytvalues. Conversely, in the alkaline side of pH_cyt K⁺ effluxseemed linked only to the E_m values. These results are consistentwith the hypothesis that the decrease in K⁺ efflux observedin non-stomatal tissues in the presence of ABA may be mediatedby the cytoplasmic acidification induced by the hormone. (Received August 6, 1996; Accepted January 19, 1997)     

Effect of Abscisic Acid on the K+ Efflux and Membrane Potential of Nicotiana tabacum L. Leaf Cells

K⁺ efflux from tobacco (Nicotiana tabacum L, cv. Samsun NN)leaf discs into the external medium was increased and the membranepotential (Em) changed in the positive direction with a changein pH from 8.0 to 4.0. E_m was affected by the external concentrationof KCl, greatly decreasing with a change in concentration from1 mM to 100 mM. The equilibrium potential of the membrane forK⁺ (E_k) was decreased in a Nernst fashion with increasing externalconcentrations of KCl. E_k is more positive than E_m above ca.50 µM KCl. Most of the experiments were carried out underconditions in which the difference between the electrochemicalpotential for K⁺ on the inside to the outside of the cell (µ_kis positive. Thus, K⁺ may passively flow to the outside of thecells accompanied by the depolarization of the membrane. Abscisic acid (ABA) increased the K⁺ efflux under conditionsof passive transport. K⁺ efflux was accelerated with an increasingconcentration of ABA, being maximal at 10^–4 M–10^–3M. This acceleration was due to the enhancement of the potassiummotive force (µ_k/F) which is the force causing the netpassive transport of K⁺. The membrane potential was decreasedfrom –205 mV to –170 mV by 2 x 10^–4 M ABAwithin 10 min. The depolarization was not transient, being lostfor at least 3 hr. These results show that ABA accelerated passive K⁺ efflux, whichaccompanied depolarization of the membrane. (Received June 22, 1981; Accepted August 24, 1981)     

Inhibition of Nitrate Assimilation in Roots in the Presence of Ammonium: The Moderating Influence of Potassium

Experiments were conducted to investigate the effect of concentrationof NH₄⁺ in nutrient solution on root assimilation of NO₃^–and to determine whether the NH₄⁺NO₃^– interaction wasmodified in the presence of K⁺. Dark-grown, detopped corn seedlings(cv. Pioneer 3369A) were exposed for 8 h to 0.15 mM Ca(NO₃)₂and varying concentrations of (NH₄)₂SO₄ in the absence or presenceof 0.15 mM K₂SO₄. The accelerated phase of NO₃^– uptakeappeared most sensitive to restriction by additions of 0.15mM (NH₄)₂SO₄. In the absence of K⁺, the restriction increasedonly slightly even when solution (NH₄)₂SO₄, was increased from0.15 mM to 12.5 mM which was accompanied by an increase of NH₄⁺in the tissue from about 7.0 to 35 µmol g^–1 fr.wt. of root. Increasing concentrations of solution NH₄⁺ progressivelyinhibited net K+ uptake. At the highest solution NH₄⁺ concentrations,there was an initial net efflux of K⁺ and no net influx occurredduring the treatment period. The severity of the NH₄)SO₄ restrictionof NO₃^– uptake was moderated considerably in the presenceof K⁺ as long as a net influx of K⁺ occurred. However, net influxof K⁺ was not associated with alteration of NH₄⁺ uptake, assimilation,or accumulation in the root tissue. The lack of correlationbetween the severity of restriction of NO₃^– uptake andendogenous NHJ suggested the restriction resulted from an effectexerted by exogenous NH₄⁺ which tended to saturate at lowersolution NHJ concentrations or by inhibitory factors generatedduring assimilation of NH₄⁺. Several mechanisms were postulatedto account for the moderating influence of K⁺. In all experiments,root NO₃^– reduction was restricted by the presence ofambient NH₄⁺. The quantitative decreases in reduction tendedto be less than decreases in NO₃^– uptake and therefore,could result from inhibition solely of uptake with subsequentlimitation in availability of substrate for the reduction process,but the possibility of a direct effect on reduction could notbe excluded.     

Flexible Coupling of Phosphate Uptake in Dunaliella acidophila at Extremely Low pH Values

The acidophilic alga Dunaliella acidophila exhibits optimalgrowth at pH 1. We have investigated the regulation of phosphateuptake by this alga using tracer techniques and by performingintracellular phosphate measurements under different growthconditions including phosphate limitation. In batch culturewith 2·2 mol m^–3 phosphate in the medium the uptakeof phosphate at micromolar phosphate concentrations followeda linear time dependence in the range of minutes and rates werein the range of 1 µmol phosphate mg^–1 chl h^–1,only. However, under discontinuous phosphate-limited growthconditions, tracer influx revealed a biphasic pattern at micromolarphosphate concentrations: An initial burst phase resulted ina 10⁴-fold internal phosphate accumulation and levelled offafter about 10 s. A double reciprocal plot of the initial influxrates obtained for phosphate-limited and unlimited algae exhibitedMichaelis-Menten kinetics. Phosphate limitation caused a significantactivation of the maximum velocity of uptake, yielding V_maxup to 1 mmol mg^–1 chl h^–1 as compared to valuesin the order of 50 µmol phosphate mg^–1 chl h^–1for the second phase (this magnitude is also representativefor non-limited batch cultures). Concomitantly the Michaelisconstant was altered from 4 mmol m^–3 to 0·7 mmolm^–3. The rapid uptake of phosphate was inhibited by arsenateand FCCP and was not stimulated by Na⁺. The pH dependence oftracer accumulation and measurements of the intracellular phosphatepool under different growth conditions indicate that at lowpH and low external phosphate concentrations the high protongradient present under these conditions is utilized for a H₃PO₄uptake or a H⁺/H₂PO^–4 cotransport. However, when the externalphosphate concentration was increased to levels sufficientlyhigh for transport to be driven by the positive membrane potential(10 mol m^–3 phosphate), the pH dependence of phosphateuptake was more complex, but could be explained by the uptakeof H₃PO₄ or a H⁺/H₂PO^–₄-cotransport at low pH and a differenttype H₂PO^–4-transport (with unknown type of ion coupling)at high pH-values. It is suggested that this flexible couplingof phosphate transport is of essential importance for the acidresistance of Dunaliella acidophila. Key words: Acid resistance, Dunaliella acidophila, phosphate cotransport, phosphate limitation, plasma membrane, sodium     

Studies on Mechano-Perception in Characeae: Effects of External Ca2+ and Cl-

The effects of modification of extracellular concentrationsof Ca²⁺ and C^– on mechano-perception were studied in internodalcells of Chara corallina. Cells were stimulated by droppinga piece of glass tubing on them, and the resulting receptorpotentials and action potentials were analyzed. When the Ca²⁺concentration was extremely lowered by adding EGTA, the amplitudesof both receptor potentials and action potentials were attenuated,suggesting the involvement of Ca²⁺ channels. However, the possibilityremained that attenuation of the amplitude of the receptor potentialwas caused by modification of membrane characteristics by extremelowering of [Ca²⁺]_o. When the plasma membrane was depolarizedto about 0 mV by adding 100 mM KC1, responses in the negativedirection were induced upon mechanical stimulation. When theplasma membrane was depolarized by adding 50 mM K²SO⁴, responsesin the positive direction were induced. Thus, Cl^– channelsmay be involved in responses induced by mechanical stimulationunder K+-induced depolarization. (Received January 16, 1996; Accepted March 25, 1997)     

K+ channel KVLQT1 located in the basolateral membrane of distal colonic epithelium is not essential for activating Cl- secretion

The cellular mechanism for Cl^– and K⁺ secretion in the colonic epithelium requires K⁺ channels in the basolateral and apical membranes. Colonic mucosa from guinea pig and rat were fixed, sectioned, and then probed with antibodies to the K⁺ channel proteins K_VLQT1 (Kcnq1) and minK-related peptide 2 (MiRP2, Kcne3). Immunofluorescence labeling for Kcnq1 was most prominent in the lateral membrane of crypt cells in rat colon. The guinea pig distal colon had distinct lateral membrane immunoreactivity for Kcnq1 in crypt and surface cells. In addition, Kcne3, an auxiliary subunit for Kcnq1, was detected in the lateral membrane of crypt and surface cells in guinea pig distal colon. Transepithelial short-circuit current (I_sc) and transepithelial conductance (G_t) were measured for colonic mucosa during secretory activation by epinephrine (EPI), prostaglandin E₂ (PGE₂), and carbachol (CCh). HMR1556 (10 µM), an inhibitor of Kcnq1 channels (Gerlach U, Brendel J, Lang HJ, Paulus EF, Weidmann K, Brüggemann A, Busch A, Suessbrich H, Bleich M, and Greger R. J Med Chem 44: 3831–3837, 2001), partially (50%) inhibited Cl^– secretory I_sc and G_t activated by PGE₂ and CCh in rat colon with an IC₅₀ of 55 nM, but in guinea pig distal colon Cl^– secretory I_sc and G_t were unaltered. EPI-activated K⁺-secretory I_sc and G_t also were essentially unaltered by HMR1556 in both rat and guinea pig colon. Although immunofluorescence labeling with a Kcnq1 antibody supported the basolateral membrane presence in colonic epithelium of the guinea pig as well as the rat, the Kcnq1 K⁺ channel is not an essential component for producing Cl^– secretion. Other K⁺ channels present in the basolateral membrane presumably must also contribute directly to the K⁺ conductance necessary for K⁺ exit during activation of Cl^– secretion in the colonic mucosa. HMR1556; K⁺ secretion; epinephrine; prostaglandin E₂; cholinergic     

Uptake of Imidazole and its Effects on the Intracellular pH and Ionic Relations of Chara corallina

Ammonia (pK_a 9.25) and methylamine (pK_a, 10.65) increase cytoplasmicpH and stimulate Cl^– influx in Chara corallina, theseeffects being associated with influx of the amine cations ona specific porter. The weak base imidazole (pK_a 6.96) has similareffects but diffuses passively into the cell both as an unionizedbase and as a cation. When the external pH is greater than 6.0influx of the unionized species predominates. Imidazole accumulates to high concentrations in the vacuole,where it is protonated. Cytoplasmic pH and vacuolar pH riseby only 0.2–0.3 units, suggesting a large balancing protoninflux across the plasma membrane. Balance of electric chargeis partially maintained by net efflux of K⁺ and net influx ofCl^–. Calculation of vacuolar concentrations of imidazole(from (¹⁴C] imidazole uptake, assuming that there is no metabolism)plus K⁺ and Na⁺ indicates an excess of cations over inorganicanions (Cl^–). However, although the osmotic potentialof the cells increases, also indicating increased solute concentrations,the increase is less than that predicted by the calculated ionicconcentrations. This discrepancy remains to be resolved. Becausethe osmotic potential also increases when imidazole is absorbedfrom Cl^–-free solutions it is likely that maintenanceof charge-balance can also involve synthesis and vacuolar storageof organic or amino acids. Key words: Imidazole, potassium, intracellular pH, membrane transport, Chara     

Interactions of NH4+ and L-glutamate with NO3- transport processes of non-mycorrhizal Fagus sylvatica roots

The processes of NO₃^– uptake and transport and the effectsof NH₄⁺ or L-glutamate on these processes were investigatedwith excised non-mycorrhizal beech (Fagus sylvatica L.) roots.NO₃^– net uptake followed uniphasic Michaelis-Menten kineticsin a concentration range of 10µM to 1 mM with an apparentK_m of 9.2 µM and a V_max of 366 nmol g^–1 FW h^–1.NH₄⁺, when present in excess to NO₃^–, or 10 mM L-glutamateinhibited the net uptake of NO₃^– Apparently, part of NO₃^–taken up was loaded into the xylem. Relative xylem loading ofNO₃^– ranged from 3.21.6 to 6.45.1% of NO₃^– netuptake. It was not affected by treatment with NH₄⁺ or L-glutamate.¹⁶N/¹³N double labelling experiments showed that NO₃^–efflux from roots increased with increasing influx of NO₃^–and, therefore, declined if influx was reduced by NH₄⁺ or L-glutamateexposure. From these results it is concluded that NO₃^–net uptake by non-mycorrhizal beech roots is reduced by NH₄⁺or L-glutamate at the level of influx and not at the level ofefflux. Key words: Nitrate transport, net uptake, influx, efflux, ammonium, Fagus, Fagaceae