Volume-regulatory responses of Amphiuma red cells in anisotonic media. The effect of amiloride

Amphiuma red cells were incubated for several hours in hypotonic or hypertonic media. They regulate their volume in both media by using ouabain-insensitive salt transport mechanisms. After initially enlarging osmotically, cells in hypotonic media return toward their original size by losing K, Cl, and H2O. During this volume-regulatory decrease (VRD) response, K loss results from a greater than 10-fold increase in K efflux. Cells in hypertonic media initially shrink osmotically, but then return toward their original volume by gaining Na, Cl, and H2O. The volume-regulatory increase (VRI) response involves a large (greater than 100-fold) increase in Na uptake that is entirely blocked by the diuretic amiloride (10(-3) M). Na transport in the VRI response shares many of the characteristics of amiloride-sensitive transport in epithelia: (a) amiloride inhibition is reversible; (b) removal of amiloride from cells pretreated with amiloride enhances Na uptake relative to untreated controls; (c) amiloride appears to act as a competitive inhibitor (Ki = 1-3 microM) of Na uptake; (d) Na uptake is a saturable function of external Na (Km approximately 29 mM); (e) Li can substitute for Na but K cannot. Anomalous Na/K pump behavior is observed in both the VRD and the VRI responses. In the VRD response, pump activity increases 3-fold despite a decrease in intracellular Na concentration, while in the VRI response, a 10-fold increase in pump activity is observed when only a doubling is predicted from increases in intracellular Na.     

Kinetics and Block of Dopamine Uptake in Synaptosomes from Rat Caudate Nucleus

The dopamine (DA) uptake system in mammalian nerve terminals was studied by measuring the unidirectional influx of tritiated DA into synaptosomes prepared from rat caudate nucleus. Two distinct time-dependent components of DA uptake were observed. The principal component was saturable with respect to DA concentration, required both external Na and Cl, and was competitively blocked by micromolar concentrations of the psychotropic agents cocaine, benztropine, nomifensine, amphetamine, and methamphetamine. This principal component of uptake has the properties expected for a carrier-mediated transport system. The second component, which accounted for about 10-30% of the DA uptake at 2 microM DA, was not saturable, and was independent of external Na, Cl, and blockers of the carrier-mediated system. The saturable, Na-dependent component had an apparent Km(DA) of about 0.5 microM. The dependence of DA uptake on external Na was sigmoid [Hill coefficient = 2; Ka(Na) = 45 mM] whereas the dependence on Cl was best described by a rectangular hyperbola [Ka(Cl) = 15 mM]. Depolarizing conditions (elevated external K) reduced the rate of DA influx. The data are consistent with a carrier-mediated DA transport mechanism in which each DA molecule entering the nerve terminal via the carrier is accompanied by two or more Na ions and one Cl ion in a rheogenic process carrying one or more net positive charges into the cell. Net, concentrative accumulation of DA inside nerve terminals may be accomplished by utilizing the Na electrochemical gradient to drive DA against its electrochemical gradient via this carrier system.     

Na(+) and Cl(-) transport by the urinary bladder of the freshwater rainbow trout (Oncorhynchus mykiss)

Freshwater (FW) rainbow trout (Oncorhynchus mykiss) urinary bladders mounted in vitro under symmetrical saline conditions displayed electroneutral active absorption of Na(+) and Cl(-) from the mucosal side; the transepithelial potential (V(t)) was 0.1 mV, and the short-circuit current was less than 1 microA cm(-2). Removal of Na(+) from mucosal saline decreased Cl(-) absorption by 56% and removal of Cl(-) decreased Na(+) absorption by 69%. However, active net absorption of both Na(+) and Cl(-) was not abolished when Cl(-) or Na(+) was replaced with an impermeant ion (gluconate or choline, respectively). Under physiological conditions with artificial urine (?Na(+) = 2.12 mM, ?Cl(-) = 3.51 mM) bathing the mucosal surface and saline bathing the serosal surface, transepithelial potential (V(t)) increased to a serosal positive approximately +7.6 mV. Unidirectional influx rates of both Na(+) and Cl(-) were 10-20-fold lower but active absorption of both ions still occurred according to the Ussing flux ratio criterion. Replacement of Na(+) with choline, or Cl(-) with gluconate, in the mucosal artificial urine yielded no change in unidirectional influx of Cl(-) or Na(+), respectively. However, kinetic analyses indicated a decrease in maximum Na(+) transport rate (J(max)) of 66% with no change in affinity (K(m)) in the low Cl(-) mucosal solution relative to the control solution. Similarly, there was a 79% decrease in J(max) values for Cl(-), again with no change in K(m), in the low-Na(+) mucosal bathing. The mucosal addition of DIDS, amiloride or bumetanide (10(-4) M) had no effect on either Na(+) or Cl(-) transport, under either symmetrical saline or artificial urine/saline conditions. Addition of the three drugs simultaneously (10(-4) M), or chlorothiazide (10(-3) M), under symmetrical saline conditions also had no effect on Na(+) or Cl(-) transport rates. Cyanide (10(-3) M) addition to mucosal artificial urine caused a slowly developing decrease of Na(+) influx to 59% and Cl(-) influx to 50% in the period after drug addition. Na(+) and Cl(-) reabsorption appears to be a partially coupled process in the urinary bladder of O. mykiss; transport mechanisms are both dependent upon and independent of the other ion.     

Stoichiometry of a half-turnover of band 3, the chloride transport protein of human erythrocytes

The kinetics of human red blood cell Cl transport have been studied under nonequilibrium conditions to determine whether or not an outward Cl gradient can recruit the transport protein from an inward-facing to an outward-facing configuration. Three kinds of evidence are consistent with this outward recruitment. First, the initial net Cl efflux into a Cl-free phosphate medium is independent of the intracellular Cl concentration in the range 20-170 mM. Second, an outward Cl gradient strongly enhances the inhibitory potency of DNDS (4,4'-dinitro-2,2'-stilbene disulfonate), which suggests that DNDS binds primarily to outward-facing states. Finally, we have estimated the number of Cl ions transported during the putative outward recruitment. Resealed red cell ghosts containing only 70 muM 36Cl were resuspended at 0 degrees C in a Cl-free, HCO3-free Na2SO4 medium. In the first 10 s, or approximately 10(6) Cl ions per ghost, followed by a much slower further loss of Cl. The rapid loss of 10(6) Cl ions per ghost, which is abolished by pretreatment with DIDS (4,4'-diisothiocyano-2,2'-stilbene disulfonate), appears to represent the Cl that is transported during the first half-turnover of the transport cycle. These data are strong evidence that the influx and efflux events in the catalytic cycle for anion transport do not take place simultaneously, and that the stoichiometry of the transport cycle is close to one pair of anions exchanged per band 3 monomer.     

Sulphate uptake and metabolism in the chrysomonad, monochrysis lutheri.

The intracellular concentration of inorganic 35SO4 in Monochrysis lutheri cells exposed to 0.513 mM Na235SO4 for up to 6-hr remained constant at about 0.038 mM. The exchange rate of this 35SO4 with the external unlabelled sulphate was negligible compared to the rate of influx across the plasmalemma (0.032 mu moles/g cells/hr). The flux of free 35SO4 to organic 35S was 0.029 mu moles/g cells/hr. Assuming an internal electrical potential in the cells of -70 mV, this intracellular concentration of inorganic 35SO4 was well in excess of that obtainable by passive diffusion as calculated from the Nernst equation. These results indicate that sulphate is accumulated by an active mechanism rather than by facilitated diffusion. Sulphate uptake appears to occur via a carrier-mediated membrane transport system which conforms to Michaelis-Menten type saturation kinetics with a Km of 3.2 X 10(-5) M and Vmax of 7.9 X 10(-5) mu moles sulphate/hr/10(5) cells. Uptake was dependent on a source of energy since the metabolic inhibitor CCCP almost completely inhibited uptake under both light and dark conditions and DCMU caused a 50% decrease in uptake under light conditions. Under dark conditions, uptake remained at about 80% of that observed under light conditions and was little affected by DCMU, indicating that the energy for uptake could be supplied by either photosynthesis or respiration. A charge and size recognition site in the cell is implied by the finding that sulphate uptake was inhibited by chromate and selenate but not by tungstate, molybdate, nitrate or phosphate. Chromate did not inhibit photosynthesis. Cysteine and methionine added to the culture medium were apparently capable of exerting inhibition of sulphate uptake in both unstarved and sulphate-starved cells. Cycloheximide slightly inhibited sulphate uptake over an 8-hr period indicating, either a slow rate of entry of the inhibitor into the cells or a slow turnover of the protein(s) associated with sulphate transport.     

Proton pump activity is required for active uptake of chloride in isolated amphibian skin exposed to freshwater

Net proton secretion and unidirectional chloride fluxes were measured in isolated skin of toads ( Bufo bufo) and frogs ( Rana esculenta) mounted in an Ussing chamber and exposed to a Ringer's solution on the serosal side and a freshwater-like solution (1-3 mM Cl(-)) on the external side. Active proton secretion was 34.2+/-2.0 pmol.cm(-2).s(-1) ( n=18) in frog skin, and 16.7+/-1.7 pmol.cm(-2).s(-1) ( n=10) in toad skin. Proton secretion by toad skin was dependent on the transepithelial potential ( V(T)), and an amiloride-insensitive short-circuit current was stimulated by exogenous CO(2)/HCO(3)(-), indicating the presence of a rheogenic proton pump. Cl(-) influx was 37.4+/-7.5 pmol.cm(-2).s(-1) ( n=14) in frog skin and 19.5+/-3.5 pmol.cm(-2).s(-1) ( n=11) in toad skin. In toad skin, the mean Cl(-) flux ratio was larger than expected for simple electro-diffusion. In 8 of 11 sets of paired skins, influx was greater than the efflux indicating active uptake of Cl(-). Cl(-) influx in toad skin was unaffected by large perturbations (100-150 mV) of V(T), which was accomplished by adding amiloride to the outer bath under open circuit conditions. A component of the Cl(-) efflux seemed to be dependent on V(T). 4,4'-Diisothiocyanato-stilbene-2,2'-disulfonic acid (DIDS; 0.3 mM or 1.3 mM) inhibited Cl(-) influx and, surprisingly, increased Cl(-) efflux in toad skin. Influx and efflux of Cl(-) in toad skin were highly dependent on the external [Cl(-)] in the freshwater range (0.1-4 mM). (36)Cl(-) influx decreased whereas the total Cl(-) efflux increased as a function of external [Cl(-)]. These data indicate the presence of a DIDS-sensitive, electroneutral carrier mechanism with an external binding site for Cl(-). Ethoxzolamide (100 micro M), an inhibitor of carbonic anhydrase, reduced proton secretion and Cl(-) influx in frog skin. Concanamycin A (0.1-10 micro M), a specific vacuolar-type proton pump (V-ATPase) inhibitor, significantly reduced proton secretion in frog skin. In addition, concanamycin A (1 micro M) significantly reduced Cl(-) influx in frog skin. We suggest that the active proton secretion and Cl(-) influx are coupled. We hypothesise that an apical V-ATPase is capable of energising active Cl(-) uptake in fresh water by creating a favourable gradient for an apical HCO(3)(-) exit in exchange for external Cl(-). The data also suggest that a carbonic anhydrase activity provides H(+) and HCO(3)(-) for apically co-expressed proton pumps and Cl(-)/HCO(3)(-) exchangers.     

Cadmium stimulation of Na-independent organic acid transport in the kidney tubules

The influence of Cd++ (as well as of Hg++ and Cu++) on the uptake of an organic acid (fluorescein) in superficial proximal tubules of the surviving rat kidney was studied at 20 degrees C, when the active transport of fluorescein does not depend on the external Na. In contrast to mercury and copper, cadmium stimulated the uptake of fluorescein from the beginning of incubation. The minimal effective concentration of Cd++ was 5 X 10(-6)M, the relative effect of Cd++ on the uptake being the same within the concentration range from 5 X 10(-6) to 10(-3) M. A 60 minutes pre-incubation with Cd++ at 20 degrees C resulted in a significant increase in the stimulatory effect of acetate on the fluorescein transport. The stimulation of the fluorescein transport by cadmium was prevented by ouabain or by omissing Na from the incubating medium, although neither ouabain nor the absence of Na affected the transport of fluorescein under these conditions. It is supposed that the stimulation by Cd++ of the fluorescein transport may result from the activated oxidation of NAD-linked substrates due to acceleration of the active transepithelial transport of Na ions.     

Dual effect of adrenalin on sugar transport in rat diaphragm muscle

The effect of adrenalin on the membrane transport of the non-metabolized sugar, 3-methylglucose, was studied in isolated "intact" rat hemidiaphragms and related to simultaneously occurring changes in the internal levels of Na+, ATP, glucose-6-P, glycerol formation and 45Ca uptake and loss. Basal sugar transport was inhibited by low (10-8-10-5 M) concentrations of adrenalin; this was antagonized by propranolol and practolol. High concentrations (10-4-10-3 M) stimulated sugar transport, and this was blocked by propranolol and butoxamine and was dependent on external Ca2+. These results suggest interaction with two different classes of adrenergic receptors, possibly of beta 1 and beta 2 types. Both low and high concentrations increases Na+ and K+ gradients by a practolol-sensitive effect. Isoproterenol behaved identically but phenylephrine had only the two practolol-sensitive effects on sugar and ion transport. Insulin did not interfere with inhibition of sugar transport and decrease in internal Na+ but prevented stimulation of sugar transport. Under anoxia adrenalin had no effect on sugar transport but led to greater Na+ gain by tissue. Addition of 3.0 mM palmitate decreased inhibition of sugar transport without changing receptor specificity. ATP was decreased and lipolysis enchanged by high adrenalin but glucose-6-P was increased by the low concentration as well. Influx of 45 Ca was decreased by low and increased by high adrenalin; 45Ca efflux was also differentially affected. The results indicate that inhibition and stimulation of sugar transport depend on different receptors and that the latter response may override the former. The data are consistent with the earlier postulated regulatory role of sarcoplasmic Ca2+ on sugar transport in muscle, with adrenalin affecting Ca2+ fluxes and distribution both directly and indirectly.     

The role of chloride in taurine transport across the human placental brush-border membrane.

Taurine, a sulfated beta-amino acid, is conditionally essential during development. A maternal supply of taurine is necessary for normal fetal growth and neurologic development, suggesting the importance of efficient placental transfer. Uptake by the brush-border membrane (BBM) in several other tissues has been shown to be via a selective Na(+)-dependent carrier mechanism which also has a specific anion requirement. Using BBM vesicles purified from the human placenta, we have confirmed the presence of Na(+)-dependent, carrier-mediated taurine transport with an apparent Km of 4.00 +/- 0.22 microM and a Vmax of 11.72-0.36 pmol mg-1 protein 20 s-1. Anion dependence was examined under voltage-clamped conditions, in order to minimize the contribution of membrane potential to transport. Uptake was significantly reduced when anions such as thiocyanate, gluconate, or nitrate were substituted for Cl-. In addition, a Cl(-)-gradient alone (under Na(+)-equilibrated conditions) could energize uphill transport as evidenced by accelerated uptake (3.13 +/- 0.8 pmol mg-1 protein 20 s-1) and an overshoot compared to Na+, Cl- equilibrated conditions (0.60 +/- 0.06 pmol mg-1 protein 20 s-1). A Cl(-)-gradient (Na(+)-equilibrated) also stimulated uptake of [3H]taurine against its concentration gradient. Analysis of uptake in the presence of varying concentrations of external Cl- suggested that 1 Cl- ion is involved in Na+/taurine cotransport. We conclude that Na(+)-dependent taurine uptake in the placental BBM has a selective anion requirement for optimum transport. This process is electrogenic and involves a stoichiometry of 2:1:1 for Na+/Cl-/taurine symport.     

Salt stress-induced chloride flux: a study using transgenic Arabidopsis expressing a fluorescent anion probe

Salt stress leads to massive accumulation of toxic levels of Na(+) and Cl(-) ions in plants. By using the recombinant fluorescent probe CLOMELEON, we demonstrate passive anion flux under salt stress. Chloride influx is restricted in the presence of divalent cations like Mg(2+) and Ca(2+), and completely blocked by La(3+). The amount but not the rate of the reported chloride uptake is independent from the kind of corresponding permeable cation (K(+) versus Na(+)), external pH and magnitude of osmotic stress. Cl(-) efflux however seems to involve stretch-activated transport. From the influence of Ca(2+) on reported changes of cytosolic anion concentrations, we speculate that transport mechanisms of Cl(-) and Na(+) might be thermodynamically coupled under saline conditions.     

Limiting CO2 levels induce a blue light-dependent HCO3- uptake system in Monoraphidium braunii

The in situ photoactivation of an HCO3- uptake system in the green alga Monoraphidium braunii requires the irradiation of the cell suspensions with short wavelength radiation (blue, UVA and/or UVC). Plasma membrane ATPase inhibitors block the uptake of this monovalent anion at pH 9. M. braunii cells grown in high CO2 lack an HCO3- uptake system in their plasma membrane, but those grown in low CO2 can take up this anion at high rates. Cells grown in high CO2, transferred to CO2-limiting conditions in the light, start taking up HCO3- in 30 min, although they take 90 min to reach maximum rates of HCO3- transport. Therefore, this induction process seems to be triggered by low external CO2 concentration. In fact, increasing or decreasing the external HCO3- concentration does not induce the uptake system and only a decrease in CO2 concentration in the medium triggers the induction process. The appearance of the HCO3- transport activity is sensitive to cycloheximide, indicating that cytoplasmic protein biosynthesis is necessary for the induction of the uptake system. Photosynthetically active radiation, but not particularly blue light, is essential for induction of the uptake system to occur and the inhibition of photosynthesis by DCMU blocks it. From these results it can be inferred that when M. braunii cells detect a drop in CO2 concentration, they induce a blue light-dependent HCO3- uptake system.     

Prolactin receptors in cultured rat mammary tumor cells. Energy-dependent uptake and degradation of hormone receptors

After energy depletion by uncouplers of oxidative phosphorylation or inhibitors of electron transport, primary cultures of carcinogen-induced rat mammary tumors have a 2- to 20-fold increase in the number of cell surface prolactin receptors. When energy-depleted cells were treated with 0.15 M NaCl plus 50 mM glycine pH 3, for 1 min at 4 degrees C, 75% of the specific surface-bound 125I-labeled ovine prolactin was removed, but prolactin and its receptor were not destroyed. Using this technique, we found that receptor-bound prolactin can be internalized (becomes resistant to pH 3.0 treatment) and then degraded. The internalization of occupied receptors required energy, was completed 30-60 min before degradation, and was independent of protein synthesis. Hormone degradation (t1/2, 42 min) but not uptake was prevented by NH4Cl or lysosomotropic amines. In the presence of cycloheximide, receptors were lost (t1/2, 62 min) unless such loss was prevented by KCN. After unoccupied receptors were activated by energy depletion, surface receptors were lost when inhibitor was removed and glucose was added. Thus, both occupied and unoccupied prolactin receptors are constantly removed from the cell surface via an energy-dependent uptake mechanism. If the receptor levels are first increased by energy depletion (with or without bound ligand) or if protein synthesis is inhibited, there is a net loss of surface binding sites. Since the receptors reappeared with 15 h after cycloheximide removal, some of the receptors probably are recycled under normal steady state conditions.     

Effect of anaerobiosis, dinitrophenol and fluoride on the active intestinal transport of galactose in snail.

The active transport of galactose across the intestinal wall (everted sacs) of the snail Cryptomphalus hortensis Müller has been studied in vitro, under several metabolic conditions. Anaerobiosis does not change the serosal/mucosal galactose gradients which are developed in oxygen atmosphere. Dinitrophenol (10(-4) M) greatly increased the O2 uptake by the tissue and clearly inhibits the sugar transport. At 5 times 10(-4) M concentration, DNP totally prevents the uphill transport while the O2 uptake is normal. The inhibition produced by DNP does not increase by anaerobiosis. Fluoride inhibits the galactose transport and also the O2 uptake. It is deduced that in snail intestine the energy for the active transport of galactose can be supplied by aerobic as much as by anaerobic metabolism. The inhibition by dinitrophenol seems to be independent of its uncoupling action on the oxidative phosphorylation. The inhibitory effect of NaF may be due both to glycolisis inhibition and to alteration of the digestive epithelium.     

Insulin and epidermal growth factor. Human fibroblast receptors related to deoxyribonucleic acid synthesis and amino acid uptake.

Receptors for insulin and epidermal growth factor (EGF) have been studied in confluent cultured intact human fibroblast monolayers. 125-I-EGF binds specifically to fibroblast monolayers. Half-maximal binding is observed at 4 times 10 minus 10 M EGF; at saturation of binding approximately 4 times 10-4 molecules of EGF are bound per cell. 125-I-Insulin is also bound specifically by intact monolayers with half-maximal binding observed at 10 minus 9 M insulin; about 4 times 10-3 molecules of insulin are bound per cell at saturation. Both insulin and EGF stimulate thymidine incorporation and alpha-aminoisobutyrate uptake. A half-maximal effect for insulin is observed at about 10 minus 9 M, both for the stimulation of thymidine incorporation and for the stimulation of alpha-aminoisobutyrate uptake; for EGF, half-maximal stimulation of both thymidine incorporation and alpha-aminoisobutyrate uptake is observed at 10 minus 10 M EGF. EGF causes an apparent greater stimulation of thymidine incorporation than does insulin, whereas the stimulation of alpha-aminoisobutyrate uptake is the same for both insulin and EGF. The degree of stimulation of alpha-aminoisobutyrate uptake by either insulin or EGF varied (1.2- to 2-fold) from one batch of cells to another, as did the measured values of the apparent K-m (average value 1 mM, range 0.6 to 2 mM) and V-max (average, 0.82, range 0.78 to 0.87 nmol/100 mug of protein per min) for alpha-aminoisobutyrate. Nonetheless, the apparent K-m of each peptide for stimulation of alpha-aminoisobutyrate uptake was independent of the degree of increase in alpha-aminoisobutyrate uptake, and was constant from one batch of cells to another. The peptide-mediated stimulation of alpha-aminoisobutyrate uptake can be attributed to a decrease in the apparent K-m for alpha-aminoisobutyrate (e.g. for insulin) from 0.70 to 0.57 mM; for EGF from 0.87 to 0.66 mM) and a concomitant increase in the apparent V-max for alpha-aminoisobutyrate (e.g. for insulin from 0.78 to 0.87 and for EGF from 0.80 to 0.84 nmol/min/100 mug of cell protein). The stimulation requires a 40- to 60-min period of preincubation with either peptide and is blocked by pretreating cells with cycloheximide. In the presence of ouabain, both peptides inhibit rather than stimulate alpha-aminoisobutyrate uptake; ouabain lowers the basal rate of alpha-aminoisobutyrate uptake. The uptake of 3-0-methyl-D-glucose is not affected by either EGF or insulin under conditions where insulin stimulates fat cell transport. These observations indicate that cultured human fibroblasts possess specific binding sites for insulin and EGF, which sites can be related to two actions of the peptides: stimulation of thymidine incorporation and alpha-aminoisobutyrate uptake.     

Phosphate transport in fertilized sea urchin eggs. II. Effects of metabolic inhibitors and studies on differentiation

Metabolic inhibitors were applied after the transport system was fully developed in concentrations sufficient to block cleavage. 0.5–1.0 × 10^?4 M cyanide and anaerobiosis caused from negligible to moderate (40%) inhibition of phosphate uptake. The inhibition occurred late in the breeding season, and the inhibitory action of cyanide on uptake was associated with irreversible developmental effects. Azide (3 × 10^?3 M) did not inhibit uptake when the chamber method was used, but the aliquot and Hopkins' tube methods gave considerable inhibition. Purified preparations of 2,4-dinitrophenol (1 × 10^?4 M) did not inhibit uptake. Sodium iodoacetate (up to 0.05 M) and phlorizin (0.005 M) exerted no effect. Calculations of the minimal work requirement for the transport process reveal that this amounts to only a small fraction (0.24% at an external phosphate concentration of 2 μM) of the total available metabolic energy. Exposure of eggs at five minutes after insemination (lag phase) to cyanide (5 × 10^?5 M), anaerobic conditions, or azide (3 × 10^?3 M) blocked the expected increase of phosphate uptake. Removal of the inhibitors led to resumption of development and the appearance of the phosphate transport system in an essentially normal pattern. Exposure of eggs to 1.4–2.0 × 10^?4 M p-chloromercuribenzoate (p-CMB) during the accumulation phase severely depressed phosphate uptake, but cleavage was not inhibited nor delayed; recovery from the inhibition was accelerated by 1 × 10^?3 M cysteine. Exposure to p-CMB during the lag phase blocked the appearance of the transport system; cleavage proceeded normally. After the removal of p-CMB little reversal occurred until the addtion of 1 × 10^?3 M cysteine, when the phosphate transport system developed in an essentially normal manner. Trypsin (0.001–0.01%) neither activates the transport system in unfertilized eggs, nor inactivates it in denuded fertilized eggs by removal of surface proteins. The data are consistent with the conclusion that (1) the phosphate transport system is newly synthesized at fertilization in energy dependent reactions, and (2) phosphate transport is a carrier mediated process not directly dependent on metabolic energy.     

Actions and interactions of norepinephrine and acetylcholine on sinus node potassium movements

The influence of norepinephrine and/or acetylcholine on K uptake of the guinea pig sinus node was investigated. Both "high" (10(-6) M) and "low" (10(-9) M) mediator concentrations were employed. The following results were obtained: 1) high concentration of norepinephrine increased K uptake; 2) high concentration of acetylcholine had a similar effect; 3) simultaneous exposure to both mediators did not lead to summation of the effects; 4) low concentration of norepinephrine depressed K uptake; 5) high concentration of acetylcholine overcame the inhibitory effect of the small concentration of norepinephrine; 6) 2-deoxy-D-glucose did not abolish the stimulating effect of acetylcholine on potassium uptake; and 7) strophanthidin abolished the stimulatory effect of norepinephrine, but not that of acetylcholine. The following conclusions are drawn: 1) both neuromediators in suitable concentrations increase K uptake in the sinus node; 2) the mechanism of such an increase is different, norepinephrine increasing the active and acetylcholine increasing the passive potassium transport; 3) a low concentration of norepinephrine depresses K uptake; and 4) when the two mediators are administered together, acetylcholine suppresses the action of norepinephrine on K transport.     

Nitrate supply affects ammonium transport in canola roots

Plants may suffer from ammonium (NH4+) toxicity when NH4+ is the sole nitrogen source. Nitrate (NO3-) is known to alleviate NH4+ toxicity, but the mechanisms are unknown. This study has evaluated possible mechanisms of NO3- alleviation of NH4+ toxicity in canola (Brassica napus L.). Dynamics of net fluxes of NH4+, H+, K+ and Ca2+ were assessed, using a non-invasive microelectrode (MIFE) technique, in plants having different NO3- supplies, after single or several subsequent increases in external NH4Cl concentration. After an increase in external NH4Cl without NO3-, NH4+ net fluxes demonstrated three distinct stages: release (tau1), return to uptake (tau2), and a decrease in uptake rate (tau3). The presence of NO3- in the bathing medium prevented the tau1 release and also resulted in slower activation of the tau3 stage. Net fluxes of Ca2+ were in the opposite direction to NH4+ net fluxes, regardless of NO3- supply. In contrast, H+ and K+ net fluxes and change in external pH were not correlated with NH4+ net fluxes. It is concluded that (i) NO3- primarily affects the NH4+ low-affinity influx system; and (ii) NH4+ transport is inversely linked to Ca2+ net flux.     

Ion uptake in Pinus banksiana treated with sodium chloride and sodium sulphate

Within its wide range across Canada, jack pine is exposed to salinity from both natural and anthropogenic sources. To compare the effects of Cl and SO₄ on salt injury, sand and solution-culture grown jack pine ( Pinus banksiana Lamb.) seedlings were treated with nutrient solutions containing 60 or 120 m M NaCl, 60 m M Na₂SO₄, or a mixture of 60 m M NaCl and 30 m M Na₂SO₄. After 5 weeks of salt treatments, concentrations of Cl, K, Na, and SO₄ were determined in roots, stem and needles of the current and previous years growth, and in necrotic needles. To determine the role of water uptake in the absorption and translocation of salts in plants, total transpiration was measured as the loss of water from a sealed system and related to total plant uptake of Cl, Na, and SO₄. Sodium uptake and root-to-shoot transport rates were greater in treatments containing Cl. A delay in root-to-shoot transport of both Na and Cl indicates retention of these ions in the roots. Electrolyte leakage of needles was more closely related to treatment Cl concentrations than treatment Na concentrations. The transport of Na ions to the shoot was related to the presence of Cl, but was not related to transpiration rate.     

Bioelectric properties and ion flow across excised human bronchi

Bioelectric properties and ion transport of excised human segmental/subsegmental bronchi were measured in specimens from 40 patients. Transepithelial electric potential difference (PD), short-circuit current (Isc), and conductance (G), averaged 5.8 mV (lumen negative), 51 microA X cm-2, and 9 mS X cm-2, respectively. Na+ was absorbed from lumen to interstitium under open- and short-circuit conditions. Cl- flows were symmetrical under short-circuit conditions. Isc was abolished by 10(-4) M ouabain. Amiloride inhibited Isc (the concentration necessary to achieve 50% of the maximal effect = 7 X 10(-7) M) and abolished net Na+ transport. PD and Isc were not reduced to zero by amiloride because a net Cl- secretion was induced that reflected a reduction in Cl- flow in the absorptive direction (Jm----sCl-). Acetylcholine (10(-4) M) induced an electrically silent, matched flow of Na+ (1.7 mueq X cm-1 X h-1) and Cl- (1.9 mueq X cm-12 X h-1) toward the lumen. This response was blocked by atropine. Phenylephrine (10(-5) M) did not affect bioelectric properties or unidirectional ion flows, whereas isoproterenol (10(-5) M) induced a small increase in Isc (10%) without changing net ion flows significantly. We conclude that 1) Na+ absorption is the major active ion transport across excised human bronchi, 2) Na+ absorption is both amiloride and ouabain sensitive, 3) Cl- secretion can be induced by inhibition of the entry of luminal Na+ into the epithelia, and 4) cholinergic more than adrenergic agents modulate basal ion flow, probably by affecting gland output.