Ion specificity of cardiac sarcolemmal Na+/H+ antiporter

In bovine cardiac sarcolemmal vesicles, an outward H+ gradient stimulated the initial rate of amiloride-sensitive uptake of 22Na+, 42K+, or 86Rb+. Release of H+ from the vesicles was stimulated by extravesicular Na+, K+, Rb+, or Li+ but not by choline or N-methylglucamine. Uptakes of Na+ and Rb+ were half-saturated at 3 mM Na+ and 3 mM Rb+, but the maximal velocity of Na+ uptake was 1.5 times that of Rb+ uptake. Na+ uptake was inhibited by extravesicular K+, Rb+, or Li+, and Rb+ uptake was inhibited by extravesicular Na+ or Li+. Amiloride-sensitive uptake of Na+ or Rb+ increased with increase in extravesicular pH and decrease in intravesicular pH. In the absence of pH gradient, there were stimulations of Na+ uptake by intravesicular Na+ and K+ and of Rb+ uptake by intravesicular Rb+ and Na+. Similarly, there were trans stimulations of Na+ and Rb+ efflux by extravesicular alkali cations. The data suggest the existence of a nonselective antiporter catalyzing either alkali cation/H+ exchange or alkali cation/alkali cation exchange. Since increasing Na+ caused complete inhibition of Rb+/H+ exchange, but saturating K+ caused partial inhibitions of Na+/H+ exchange and Na+/Na+ exchange, the presence of a Na(+)-selective antiporter is also indicated. Although both antiporters may be involved in pH homeostasis, a role of the nonselective antiporter may be in the control of Na+/K+ exchange across the cardiac sarcolemma.     

Cation-selective channels in amphibian epithelia: electrophysiological properties and activation

1. Na+ as well as Li+ move across the apical membrane through amiloride-sensitive ionic channels. 2. K+ movements across the apical membrane occur through Ba2+- and Cs+-sensitive channels which do not allow the passage of Na+ or Li+. 3. A third pathway in the apical membrane is permeable for Na+, K+, Cs+, Rb+, NH+4 and Ti+. The currents carried by these monovalent cations are blocked by Ca2+ and divalent cations as well as La3+. 4. In the urinary bladder, the Ca2+-sensitive currents are stimulated by oxytocin, activators of cytosolic cAMP and cAMP analogues. Also the oxytocin activated currents are blocked by divalent cations and La3+. 5. Nanomolar concentrations of mucosal Ag+ activate the third channel and open the pathway for movements of Ca2+, Ba2+ and Mg2+, which are known to permeate through Ca2+ channels in excitable tissues.     

Trans-stimulation and driving forces for GSH transport in sinusoidal membrane vesicles from rat liver

Sinusoidal membrane vesicles from rat liver were employed to study the characteristics of GSH transport. Saturable concentration dependent uptake was best described by the sum of a high and low Km transport. Preloading with GSH markedly stimulated the initial uptake of GSH. GSH transport was electrogenic; uptake was enhanced by an inwardly directed K+ gradient which could be blocked by the K+-channel blocker, Ba2+. The other cations such as Na+, Li+ were poor substitutes for K+. These results therefore show that net GSH transport involves movement of K+.     

Insulin stimulation of (Na+,K+)-adenosine triphosphatase-dependent 86Rb+ uptake in rat adipocytes

Insulin stimulated the uptake of 86Rb+ (a K+ analog) in rat adipocytes and increased the steady state concentration of intracellular potassium. Half-maximal stimulation occurred at an insulin concentration of 200 pM. Both basal- and insulin-stimulated 86Rb+ transport rates depended on the concentration of external K+, external Na+, and were 90% inhibited by 10(-3) M ouabain and 10(-3) M KCN, indicating that the hormone was activating the (Na+,K+)-ATPase. Insulin had no effect on the entry of 22Na+ or exit of 86Rb+. Kinetic analysis demonstrated that insulin acted by increasing the maximum velocity, Vmax, of 86Rb+ entry. Inhibition of the rate of Rb+ uptake by ouabain was best described by a biphasic inhibition curve. Scatchard analysis of ouabain binding to intact cells indicated binding sites with multiple affinities. Only the rubidium transport sites which exhibited a high affinity for ouabain were stimulated by insulin. Stimulation required insulin binding to an intact cell surface receptor, as it was reversible by trypsinization. We conclude that the uptake of 86Rb+ by the (Na+,K+)-ATPase is an insulin-sensitive membrane transport process in the fat cell.     

Energetics of alanine, lysine, and proline transport in cytoplasmic membranes of the polyphosphate-accumulating Acinetobacter johnsonii strain 210A.

Amino acid transport in right-side-out membrane vesicles of Acinetobacter johnsonii 210A was studied. L-Alanine, L-lysine, and L-proline were actively transported when a proton motive force of -76 mV was generated by the oxidation of glucose via the membrane-bound glucose dehydrogenase. Kinetic analysis of amino acid uptake at concentrations of up to 80 microM revealed the presence of a single transport system for each of these amino acids with a Kt of less than 4 microM. The mode of energy coupling to solute uptake was analyzed by imposition of artificial ion diffusion gradients. The uptake of alanine and lysine was driven by a membrane potential and a transmembrane pH gradient. In contrast, the uptake of proline was driven by a membrane potential and a transmembrane chemical gradient of sodium ions. The mechanistic stoichiometry for the solute and the coupling ion was close to unity for all three amino acids. The Na+ dependence of the proline carrier was studied in greater detail. Membrane potential-driven uptake of proline was stimulated by Na+, with a half-maximal Na+ concentration of 26 microM. At Na+ concentrations above 250 microM, proline uptake was strongly inhibited. Generation of a sodium motive force and maintenance of a low internal Na+ concentration are most likely mediated by a sodium/proton antiporter, the presence of which was suggested by the Na(+)-dependent alkalinization of the intravesicular pH in inside-out membrane vesicles. The results show that both H+ and Na+ can function as coupling ions in amino acid transport in Acinetobacter spp.     

The ATP dependence of a ouabain-sensitive sodium efflux activated by external sodium, potassium and lithium in human red cells.

The stimulation of ouabain-sensitive Na+ efflux by external Na+, K+ and Li+ was studied in control and ATP-depleted human red cells. In the presence of 5 mM Na+, with control and depleted cells, Li+ stimulated with a lower apparent affinity than K+, and gave a smaller maximal activation than K+. The ability of Na+, K+ and Li+ to activate Na+ efflux was a function of the ATP content of the cells. Relative to K+ both Na+ and Li+ became more effective activators when the ATP was reduced to about one tenth of the control values. At this low ATP concentration Na+ was absolutely more effective than K+.     

Regulation of glucose transport in Ca2+-tolerant myocytes from adult rat heart

Calcium-tolerant cardiac myocytes were isolated from adult rat ventricles and sarcolemmal glucose transport was assessed by measuring linear initial uptake rates of the nonmetabolized glucose analog 3-O-methyl-D-glucose in the presence and absence of Ca2+ in the incubation medium. (1) Agents which are known to increase internal Na+ and thus stimulate Ca2+ influx via Na+-Ca2+ exchange stimulated 3-methylglucose transport in the presence of external Ca2+. These include low-Na+ medium, 10(-6) M ouabain and K+-free medium, cyanide and the sodium ionophore, monensin. Hyperosmolarity stimulated transport also in the absence of Ca2+, consistent with release of Ca2+ from internal stores. Transport was decreased in a hypo-osmolar medium and with 10(-9) M ouabain, a concentration which stimulates the Na+ pump. (2) The calcium ionophore A23187 increased basal 3-methylglucose transport but opposed stimulation of transport by insulin. (3) Insulin-stimulated transport was antagonized by palmitate and this effect was reversed by 2-bromostearate, an inhibitor of fatty acid oxidation. These results are identical in all respects to those obtained in intact cardiac and skeletal muscle preparations, confirming that hexose transport in muscle shows Ca2+ dependence and indicating that isolated cardiac myocytes are suitable for the study of this phenomenon.     

Sodium-dependent glucose transport by cultured proximal tubule cells

The cotransport of sodium ion and alpha-methyl glucose, a non-metabolized hexose, was studied in rabbit proximal tubule cells cultured in defined medium. The rate of uptake of alpha-methyl glucose shows saturation kinetics, in which Km, but not Vmax, is dependent upon the Na+ concentration in the medium. The transport system was found to be of the high-affinity type, characteristic of the straight portion of the proximal tubule. Analysis of the rates of initial uptake within the context of a generalized cotransport model, suggests that two Na+ ions are bound in the activation of the hexose transport. The steady-state level of accumulation of alpha-methyl glucose also depends upon sodium concentration, consistent with the initial rate findings. The uptake of alpha-methyl glucose is inhibited by other sugars with the relative potencies of D-glucose greater than alpha-methyl glucose greater than D-galactose = 3-O methylglucose. L-Glucose, D-fructose, and D-mannose show no inhibition. Phlorizin inhibits the alpha-methyl glucose uptake with a Ki of 9 X 10(-6) M. Ouabain (10(-3) M) decreases the steady-state alpha-methyl glucose accumulation by 60%. In the absence of sodium, the accumulation of alpha-methyl glucose is 7-fold less than at 142 mM Na+, reaching a level comparable to the sodium-independent accumulation of 3-O-methyl-D-glucose. These findings are similar to those observed in the proximal tubule of the intact kidney.     

Distinct effects of various amino acids on 45Ca2+ fluxes in rat pancreatic islets.

The effects of three types of amino acids on 45Ca2+ fluxes in rat pancreatic islets have been compared. Alanine, a non-insulinotropic neutral amino acid, transported with Na+, increased 45Ca2+ efflux in the presence or in the absence of extracellular Ca2+, but not in the absence of Na+. Its effects in Na+-solutions were practically abolished by 7 mM-glucose. Alanine slightly stimulated 45Ca2+ influx (5 min uptake) only when Na+ was present. Two insulinotropic cationic amino acids (arginine and lysine) triggered similar changes in 45Ca2+ efflux. They accelerated the efflux in the presence of Ca2+ and inhibited the efflux in a Ca2+-free medium, whether glucose was present or not. In an Na+-free Ca2+-medium, arginine and lysine markedly accelerated 45Ca2+ efflux, but this effect was suppressed by 7 mM-glucose. Arginine stimulated 45Ca2+ influx irrespective of the presence or absence of glucose and Na+. Leucine, a neutral insulinotropic amino acid well metabolized by islet cells, inhibited 45Ca2+ efflux from the islets in a Ca2+-free medium; this effect was potentiated by glutamine. In the presence of Ca2+ and Na+, leucine was ineffective alone, but triggered a marked increase in 45Ca2+ efflux when combined with glutamine. In an Na+-free Ca2+-medium, leucine accelerated 45Ca2+ efflux to the same extent with or without glutamine. Leucine also stimulated 45Ca2+ influx in the presence or in the absence of Na+, but its effects were potentiated by glutamine only in the presence of Na+. The results show that amino acids of various types cause distinct changes in 45Ca2+ fluxes in pancreatic islets. Certain of these changes involve an Na+-mediated mobilization of cellular Ca2+ from sequestering sites where glucose appears to exert an opposite effect.     

Na+/Ca2+ antiport in cultured arterial smooth muscle cells. Inhibition by magnesium and other divalent cations

Cultured smooth muscle cells from rat aorta were loaded with Na+, and Na+/Ca2+ antiport was assayed by measuring the initial rates of 45Ca2+ influx and 22Na+ efflux, which were inhibitable by 2',4'-dimethylbenzamil. The replacement of extracellular Na+ with other monovalent ions (K+, Li+, choline, or N-methyl-D-glucamine) was essential for obtaining significant antiport activity. Mg2+ competitively inhibited 45Ca2+ influx via the antiporter (Ki = 93 +/- 7 microM). External Ca2+ or Sr2+ stimulated 22Na+ efflux as would be expected for antiport activity. Mg2+ did not stimulate 22Na+ efflux, which indicates that Mg2+ is probably not transported by the antiporter under the conditions of these experiments. Mg2+ inhibited Ca2+-stimulated 22Na+ efflux as expected from the 45Ca2+ influx data. The replacement of external N-methyl-D-glucamine with K+, but not other monovalent ions (choline, Li+), decreased the potency of Mg2+ as an inhibitor of Na+/Ca2+ antiport 6.7-fold. Other divalent cations (Co2+, Mn2+, Cd2+, Ba2+) also inhibited Na+/Ca2+ antiport activity, and high external potassium decreased the potency of each by 4.3-8.6-fold. The order of effectiveness of the divalent cations as inhibitors of Na+/Ca2+ antiport (Cd2+ greater than Mn2+ greater than Co2+ greater than Ba2+ greater than Mg2+) correlated with the closeness of the crystal ionic radius to that of Ca2+.     

Modulation of serotonin uptake kinetics by ions and ion gradients in human placental brush-border membrane vesicles

The modulation of serotonin uptake kinetics by Na+, Cl-, H+, and K+ was investigated in brush-border membrane vesicles prepared from normal human term placentas. The presence of Na+ and Cl- in the external medium was mandatory for the function of the serotonin transporter. In both cases, the initial uptake rate of serotonin was a hyperbolic function of the ion concentration, indicating involvement of one Na+ and one Cl- per transport of one serotonin molecule. The apparent dissociation constant for Na+ and Cl- was 145 and 79 mM, respectively. The external Na+ increased the Vmax of the transporter and also increased the affinity of the transporter for serotonin. The external Cl- also showed similar effects on the Vmax and the Kt, but its effect on the Kt was small compared to that of Na+. The presence of an inside-acidic pH, with or without a transmembrane pH gradient, stimulated the NaCl-dependent serotonin uptake. The effect of internal [H+] on the transport function was to increase the Vmax and decrease the affinity of the transporter for serotonin. The presence of K+ inside the vesicles also greatly stimulated the initial rates of serotonin uptake, and the stimulation was greater at pH 7.5 than at pH 6.5. This stimulation was a hyperbolic function of the internal K+ concentration at both pH values, indicating involvement of one K+ per transport of one serotonin molecule. The apparent dissociation constant for K+ was 5.6 mM at pH 6.5 and 4.0 mM at pH 7.5. The effects of internal [K+] on the uptake kinetics were similar to those of internal [H+].(ABSTRACT TRUNCATED AT 250 WORDS)     

Translocation of fatty acids across the basolateral rat liver plasma membrane is driven by an active potential-sensitive sodium-dependent transport system

In previous studies it was shown that hepatocellular uptake of fatty acids is mediated by a specific fatty acid binding membrane protein. To determine now directly the driving forces for their entry into hepatocytes, the uptake of a representative long chain fatty acid, [3H]oleate, by basolateral rat liver plasma membrane vesicles was examined. Influx of oleate was stimulated by increasing the Na+ concentration of the medium. In the presence of an inwardly directed Na+ gradient (NaSCN, NaNO3, NaCl) oleate was accumulated during the initial uptake phase (20 s) at a concentration of 1.4-1.9-fold that at equilibrium (overshoot). This activation of influx was not observed after replacement of Na+ by Li+, K+, or choline+. Na+-dependent oleate uptake was significantly stimulated by creation of a negative intravesicular potential, either by altering the accompanying anions or by valinomycin-induced K+ diffusion potentials, suggesting an electrogenic transport mechanism. Na+-dependent fatty acid uptake was temperature dependent, with maximal overshoots occurring at 37 degrees C, and revealed saturation kinetics with a Km of 83.1 nM and Vmax of 2.9 nmol X min-1 X mg protein-1. These studies demonstrate that the carrier-mediated hepatocellular uptake of fatty acids represents an active potential-sensitive Na+-fatty acid cotransport system.     

Deactivation of the respiratory burst in activated macrophages: evidence for alteration of signal transduction

Enhanced function of the respiratory burst, measured as stimulated release of superoxide anion (O2-) or hydrogen peroxide, characterizes activated macrophages. Activated macrophages undergo a decline in their capacity to release O2- (a deactivation) when placed in culture for 3 days. To better understand the molecular basis for the enhanced respiratory burst of activated macrophages, we explored the mechanisms underlying deactivation of activated mouse peritoneal macrophages. Deactivation was observed when the assay was performed in a physiologic Na+ buffer, and by day 3 of culture, release of O2- from activated macrophages stimulated with phorbol myristate acetate (PMA) was almost identical to that in resident (nonactivated) macrophages. In contrast, when the assay was performed in a buffer in which Na+ was replaced by K+, release of O2- from activated macrophages on day 3 was equal to or greater than that on day 0, suggesting that the enzyme responsible for the respiratory burst was not altered during culture. The number and affinity of PMA receptors were not changed during culture and were not affected by high external K+. Continuous assay of O2- release by coverslip-adherent macrophages in a cuvette indicated that the lag time between addition of stimulus and release of O2- was reduced, and the initial rate of O2- release was enhanced in K+ buffer. The potency of monovalent cations to support O2- release was K+ greater than Rb+ greater than choline+ greater than Cs+ = Na+ greater than Li+, suggesting that characteristics such as ionic radius or molecular size influence this effect, and the effect is not due simply to absence of Na+. Extracellular Ca2+ or Mg2+ was required for the maximal effect of high external K+, and enhancement by high K+ and divalent cations increased progressively during culture. These findings suggest that deactivation is caused primarily by changes in signal transduction from PMA receptors to the respiratory burst enzyme, rather than by changes in these receptors or the enzyme itself, and that signal transduction can differ in different macrophage populations.     

Sodium gradient-dependent L-glutamate transport is localized to the canalicular domain of liver plasma membranes. Studies in rat liver sinusoidal and canalicular membrane vesicles

The driving forces for L-glutamate transport were determined in purified canalicular (cLPM) and basolateral (i.e. sinusoidal and lateral; blLPM) rat liver plasma membrane vesicles. Initial rates of L-glutamate uptake in cLPM vesicles were stimulated by a Na+ gradient (Na+o greater than Na+i), but not by a K+ gradient. Stimulation of L-glutamate uptake was specific for Na+, temperature sensitive, and independent of nonspecific binding. Sodium-dependent L-glutamate uptake into cLPM vesicles exhibited saturation kinetics with an apparent Km of 24 microM, and a Vmax of 21 pmol/mg X min at an extravesicular sodium concentration of 100 mM. Specific anionic amino acids inhibited L-[3H]glutamate uptake and accelerated the exchange diffusion of L-[3H]glutamate. An outwardly directed K+ gradient (K+i greater than K+o) further increased the Na+ gradient (Na+o greater than Na+i)-dependent uptake of L-glutamate in cLPM vesicles, resulting in a transient accumulation of L-glutamate above equilibrium values (overshoot). The K+ effect had an absolute requirement for Na+. In contrast, in blLPM the initial rates of L-glutamate uptake were only minimally stimulated by a Na+ gradient, an effect that could be accounted for by contamination of the blLPM vesicles with cLPM vesicles. These results indicate that hepatic Na+ gradient-dependent transport of L-glutamate occurs at the canalicular domain of the plasma membrane, whereas transport of L-glutamate across sinusoidal membranes results mainly from passive diffusion. These findings provide an explanation for the apparent discrepancy between the ability of various in vitro liver preparations to transport glutamate and suggest that a canalicular glutamate transport system may serve to reabsorb this amino acid from bile.     

Regulation of 22Na+ transport by calcium in an established kidney epithelial cell line.

The role of calcium in regulating the Na+ channel in an established kidney epithelial cell line has been examined. Extracellular calcium was inhibitory to Na+ uptake, and a Dixon plot of the initial Na+ uptake rate in the presence of Ca2+ was nonlinear, suggesting a mixed pattern of inhibition. Similar patterns of inhibition were also observed for other divalent cations, including Ba2+, Mg2+, and Mn2+. In contrast elevated concentrations of intracellular calcium resulted in a stimulation of Na+ entry. This intracellular effect was specific to calcium, with Mg2+ and Mn2+ appearing much less effective. Lineweaver-Burk plots of Na+ influx in calcium-loaded and unloaded cells were linear, suggesting that under both conditions a single system transported Na+. Although Na+ entry was stimulated by intracellular Ca2+, the cells did not exhibit other counter transport phenomena reported with cell types in which a Na+/Ca2+ exchange system is operative. Thus, the results indicate that calcium acts as an allosteric regulator of Na+ transport by the Na+ channel.     

Mechanism of proline transport in Escherichia coli K12. III. Inhibition of membrane potential-driven proline transport by syn-coupled ions and evidence for symmetrical transition states of the 2H+/proline symport carrier

Specific inhibition of 2H+/proline symport by syn-coupled ions (Na+, Li+, and H+) was investigated using cytoplasmic membrane vesicles prepared from the proline carrier-overproducing strain MinS/ pLC4 -45 of Escherichia coli K12. The 2H+/proline symport driven by the membrane potential generated via respiration with 20 mM ascorbate/Tris, 0.1 mM phenazine methosulfate was specifically inhibited by Na+. The inhibition by Na+ was described by a fully noncompetitive mechanism, and the apparent Ki for Na+ was 15 mM. A linear correlation between the apparent Vmax and the apparent Kd was observed. Li+ stimulated the transport activity 2-fold at 10 mM and inhibited it at concentrations above 50 mM. H+ caused fully noncompetitive inhibition of 2H+/proline symport, and its apparent Ki was 0.6 microM. These results indicate that the concentrations of Na+ and H+ strictly and independently regulate the amount of the active C state carrier responsible for 2H+/proline symport driven by the membrane potential by inhibiting the transition from the C* state carrier which exhibits Na+- and H+-dependent binding of proline and is predominant in nonenergized conditions.     

The effect of ions on the adhesion and internalization of Chlamydia trachomatis by HeLa cells

The adhesion and internalization of Chlamydia trachomatis by HeLa cells was unaffected by removal of K+, Mg2+, or glucose from the incubation medium, slightly reduced by removal of Na+, and significantly reduced by omission of Ca2+, Sr2+, Mg2+, and Mn2+ could replace Ca2+ in the adhesion but only Sr2+ supported internalization, and La3+, Co2+, Fe3+, Ba2+, and Zn2+ all reduced internalization more than adhesion. During initial infection there was no measurable difference in the uptake or release of 45Ca2+ or 86Rb+ between infected and noninfected HeLa monolayers. Infection was not prevented by pretreatment of the monolayers with the calcium channel blockers, verapamil, D600, and nitrendipine, or the calmodulin inhibitors, TMB-8 or trifluperazine. The results suggest that divalent cations are not essential for chlamydial infection but that the process of internalization is facilitated by the presence of cations, particularly Na+ and Ca2+.     

The asymmetric effect of lanthanides on Na+-gradient-dependent Ca2+ transport in synaptic plasma membrane vesicles

Lanthanides (La3+, Pr3+ and Tb3+) inhibit Na+-gradient-dependent Ca2+ influx into synaptic plasma membrane vesicles. 50% inhibition is obtained by 7 microM lanthanide concentration. The inhibition of the Na+-gradient-dependent Ca2+ uptake exhibits competitive kinetic behaviour. The apparent Km of the Ca2+ influx is increased from 50 microM in the absence of lanthanides to 118 microM in the presence of La3+, 170 microM in the presence of Pr3+ and 130 microM in the presence of Tb3+. The maximal reaction velocity is not altered (8.35 nmol Ca2+ transported per mg protein per min in the absence of lanthanides and 8.16 nmol/mg per min in the presence of lanthanides). Lanthanides also inhibited Na+-gradient-dependent Ca2+ efflux from synaptic plasma membrane vesicles that were preloaded with Ca2+ in a Na+-gradient-dependent manner. Introduction of La3+ into the interior of the synaptic plasma membrane vesicles by rapid freezing of the vesicles in liquid N2 and slow thawing had no effect on either Na+-gradient-dependent Ca2+ influx or efflux. Synaptic plasma membrane vesicles can be preloaded with Ca2+ also in an ATP-dependent manner. This form of Ca2+ uptake is also inhibited by La3+ though at higher concentrations than the Na+-gradient-dependent Ca2+ uptake. Na+-gradient-dependent efflux from synaptic plasma membrane vesicles preloaded in an ATP-dependent fashion ('inside-out' vesicles) unlike efflux from synaptic plasma membrane vesicles preloaded in a Na+-gradient-dependent manner was not inhibited by La3+. These findings suggest that the inhibition by La3+ is manifested asymmetrically on both sides of the synaptic plasma membrane. Lanthanides are probably not transported via the Na+-Ca2+ exchanger since Tb3+ entry measured by fluorescence of Tb3+-dipicolinic acid complex formation occurred at high Tb3+ concentrations only (1.5 mM or above) and was not Na+-gradient dependent.     

Effect of internally loaded iodide, thiocyanate, and perchlorate on sodium-dependent iodide uptake by phospholipid vesicles reconstituted with thyroid plasma membranes: iodide counterflow mediated by the iodide transport carrier

Na+-dependent I- transport and I- counterflow were studied using phospholipid vesicles (P-vesicles) made of porcine thyroid plasma membranes and soybean phospholipid by sonication. 1) I- uptake by P-vesicles incubated in the presence of external Na+ was higher than that by P-vesicles incubated in choline+ instead of Na+. The vesicles exhibited Na+-dependent I- uptake. When P-vesicles were internally loaded with I- prior to incubation in Na+, a further increase in Na+-dependent I- uptake was observed, although the concentration of internal I- was very much higher than that outside. In the absence of external Na+, I- uptake by P-vesicles preloaded with I- was comparable to baseline uptake. 2) Na+-dependent I- uptake by P-vesicles not loaded with I- and enhanced Na+-dependent I- uptake by P-vesicles preloaded with I- were both inhibited by either of SCN- and ClO4- added outside the vesicles. 3) When P-vesicles were loaded with SCN- instead of I- and incubated in Na+, I- uptake by these vesicles was also higher than baseline Na+-dependent I- uptake. However, a ClO4- load did not result in an increase in I- uptake. These results indicate that Na+-dependent I- transport including Na+-dependent I- counterflow is specifically mediated by the thyroid I- carrier. SCN- - I- counterflow in addition to I- - I- counterflow occurs dependently on Na+, but ClO4- - I- counterflow does not.     

22Na+ fluxes in thymic lymphocytes. I. Na+/Na+ and Na+/H+ exchange through an amiloride-insensitive pathway

The Na+ transport pathways of normal rat thymocytes were investigated. Na+ conductance was found to be lower than K+ conductance, which is consistent with reported values of membrane potential. In contrast, the isotopically measured Na+ permeability was greater than 10-fold higher than that of K+, which indicates that most of the flux is electroneutral. Cotransport with Cl- (or K+ and Cl-) and countertransport with Ca2+ were ruled out by ion substitution experiments and use of inhibitors. Countertransport for Na+ or H+ through the amiloride-sensitive antiport accounts for only 15-20% of the resting influx. In the presence of amiloride, 22Na+ uptake was increased in Na+-loaded cells, which suggests the existence of Na+/Na+ countertransport. Cytoplasmic pH determinations using fluorescent probes indicated that under certain conditions this amiloride-resistant system will also exchange Na+ for H+, as evidenced by an internal Na+- dependent acidification is proportional to internal [Na+] but inversely related to extracellular [Na+]. Moreover, 22Na+ uptake is inhibited by increasing external [H+]. The results support the existence of a substantial amiloride-insensitive, electroneutral cation exchange system capable of transporting Na+ and H+.