Cotransport Rate Research Articles

High Affinity Transport of CO2 in the Cyanobacterium Synechococcus UTEX 625

The active transport of CO(2) in Synechococcus UTEX 625 was measured by mass spectrometry under conditions that preclude HCO(3) (-) transport. The substrate concentration required to give one half the maximum rate for whole cell CO(2) transport was determined to be 0.4 +/- 0.2 micromolar (mean +/- standard deviation; n = 7) with a range between 0.2 and 0.66 micromolar. The maximum rates of CO(2) transport ranged between 400 and 735 micromoles per milligram of chlorophyll per hour with an average rate of 522 for seven experiments. This rate of transport was about three times greater than the dissolved inorganic carbon saturated rate of photosynthetic O(2) evolution observed under these conditions. The initial rate of chlorophyll a fluorescence quenching was highly correlated with the initial rate of CO(2) transport (correlation coefficient = 0.98) and could be used as an indirect method to detect CO(2) transport and calculate the substrate concentration required to give one half the maximum rate of transport. Little, if any, inhibition of CO(2) transport was caused by HCO(3) (-) or by Na(+)-dependent HCO(3) (-) transport. However, (12)CO(2) readily interfered with (13)CO(2) transport. CO(2) transport and Na(+)-dependent HCO(3) (-) transport are separate, independent processes and the high affinity CO(2) transporter is not only responsible for the initial transport of CO(2) into the cell but also for scavenging any CO(2) that may leak from the cell during ongoing photosynthesis.

Red blood cell Na+ transport as a predictor of blood pressure response to Na+ load in young blacks and whites.

The present study was designed to investigate the role of abnormalities in red blood cell sodium-potassium-chloride (Na-K-Cl) cotransport and Na+ pump as predictors of the pressor response to chronic oral Na+ loading in young whites and blacks. Subjects were healthy adults from 18 to 23 years of age and included normotensive whites (n = 24) and normotensive blacks (n = 35). Red blood cell transport studies were performed before Na+ loading. The Na+ load consisted of 10 g NaCl daily added to the usual diet. A sodium-sensitive response was defined as an increase of 5 mm Hg or more in mean arterial pressure after the Na+ load; a sodium-insensitive response was a less than 5 mm Hg increase in mean arterial pressure. A sodium-sensitive response occurred in 16% of whites and 57% of blacks. Black subjects have a significantly lower (p less than 0.005) maximal rate of furosemide-sensitive Na+ efflux and a higher Km for cellular Na+ (p less than 0.05) to activate Na-K-Cl cotransport than white subjects. Normotensive blacks with sodium-sensitive blood pressure response had a higher Km (14.4 +/- 6 mmol/l cell, n = 17, mean +/- SD) to activate the cotransport than sodium-insensitive blacks (9.9 +/- 3.7 mmol/l cell, n = 13, p less than 0.001). Normotensive whites had a significantly lower red blood cell Na+ content (p less than 0.05) and a higher maximal rate of cotransport (p less than 0.005) than young normotensive blacks.(ABSTRACT TRUNCATED AT 250 WORDS)

The Na +-dependent proline carrier, of eel intestinal brush-border membrane, sequentially binds proline and then Na +

The mechanism of Na +/ l-proline cotransport, present on brush-border membrane (BBM) vesicles of the European eel intestine, was studied. Initial cotransport rates, depending on increasing proline and Na + concentrations in the extravesicular medium (zero-trans conditions), were measured by monitoring the decay of an inside-negative membrane potential, i.e. the fluorescence quenching of the voltage-sensitive cyanine dye 3,3′-diethylthiacarbocyanine iodide (DiS-C 2(5)). By simultaneously estimating the substrate-dependent Na +-influx (with the fluorescent dye) and the Na +-dependent [ 3H]substrate influx, it was concluded that proline was cotransported with 1 Na + ion and glucose with 2 Na + ions. The kinetics of proline/Na + cotransport were then investigated. Graphical analysis excluded a ping-pong mechanism. Under rapid equilibrium assumptions, by fitting model equations to rate values it was possible to exclude the random and the ordered Na +/proline mechanisms. Therefore, in eel intestinal BBM vesicles, the mechanism of proline/Na + cotransport is ordered and proline out binds to the carrier prior to Na + out.

Renal Fanconi syndrome: developmental basis for a new animal model with relevance to human disease

Using succinylacetone (SA), a metabolite of tyrosine excreted in excess by infants and children with hereditary tyrosinemia and the renal Fanconi syndrome (FS), we have investigated developmentally-related membrane transport events leading to emergence of the generalized renal tubular dysfunction seen in human FS. SA was found to impair sugar and amino acid uptake by both newborn renal tubules and 7-day renal brush-border membrane vesicles (BBMV). This impairment by SA was due in part to a slowing of substrate cotransport rate of 22Na+-entry into BBMV. Concentration-dependent uptake studies indicated SA inhibited the newborn high-affinity transport systems for sugars and amino acids. SA also caused an increase in membrane fluidity and a shift in the thermotropic transition temperature. The demonstrated dual nature of SA's effect on membrane fluidity and O2 consumption, together with the relative contribution of each component to SA-induced transport impairment helps to provide a basis for an understanding of the age-related increases in glucosuria, aminoaciduria and natriuria seen in infants with FS.

Properties and Diversity of (Na-K-C1) Cotransporters

A specific pathway that mediates the coupled, electrically neutral transport of Na, K, and Cl ions across cell membranes is present in a wide variety of animal tissues, and appears to serve a number of different physiological functions. (Na-K-Cl) cotransport is involved in transport of salt and water across both reabsorptive and secretory epithelia, including the thick ascending limb of Henle's loop (TALH) of mammalian kidney (33), distal tubule of amphibian kidney (67), Necturus gallbladder (12), shark rectal gland (46), rabbit parotid (91), and flounder intestine (72, 73). In other cells, including Ehrlich ascites tumor cells (30, 31) and human red blood cells (17), (Na-K­ Cl) cotransport may play a role in the maintenance and regulation of cell volume. In excitable cells, such as squid axon (4, 85) and cardiac myocytes (57), this cotransporter may regulate the ion gradients that determine the resting membrane potential, and recent evidence suggests that the cotransport system of chick cardiac myocytes is stimulated by catecholamines, which enhance cardiac contractility (57). In avian red cells, (Na-K-Cl) cotransport is also stimulated by ,B-adrenergic catecholamines (44, 89, 92), and the high rate of cotransport in these cells may contribute to extrarenal potassium regulation in birds (43). Reduced levels of (Na-K-Cl) cotransport in vascular smooth muscle cells may reflect or even contribute to the pathogenesis of essential hypertension (71). As we continue to find (Na-K-Cl) cotransport in an increasing number of tissues, this pathway will likely be found to serve still other physiological functions, and abnormalities in its operation or regulation may prove to be important in the pathophysiology of specific disease states. In this review, we compare properties of (Na-K-CI) cotransport systems

Hemodialysis and red cell cation transport in uremia: Role of membrane free fatty acids

Active and facilitated cation transport in erythrocytes of uremic patients may be improved acutely by hemodialysis, although the mechanisms remain unknown. As nonesterified fatty acids (NEFA) can affect Na+ pump activity in vitro, changes in plasma and red cell membrane NEFA content following a single hemodialysis procedure were examined and compared with acute changes in erythrocyte cation flux rates in 34 hemodialysis patients. In nonsodium-loaded cells, small changes in Na+ pump flux with dialysis did correlate with changes in intracellular Na+ content (r = 0.59; N = 17; P less than 0.01). On average, neither maximal Na+ pump activity nor Na+/Li+ counter-transport flux improved with dialysis, but Na+/K+/Cl- cotransport rates rose 25% post-dialysis (P less than 0.02). Plasma NEFA levels rose 87% following hemodialysis but erythrocyte membrane NEFA content declined by 23% (P less than 0.001). Importantly, 24 of the 34 subjects studied had a decrease in erythrocyte membrane NEFA content of greater than 10%, and in these patients, the fall in membrane NEFA correlated with an increase in ouabain-sensitive Na+ efflux (r = 0.564; P less than 0.01). The effects of hemodialysis on both erythrocyte NEFA content and Na+ pump flux could be reproduced by incubating pre-dialysis cells in fatty acid-free albumin. We conclude that acute changes in membrane NEFA may modulate active cation transport in uremic erythrocytes.

Influence of race, sex, and blood pressure on erythrocyte sodium transport in humans.

Sodium transport of erythrocytes from normotensive and essential hypertensive subjects was evaluated by determining ouabain-sensitive and ouabain-insensitive sodium efflux rates, Na+-Li+ countertransport rates, Li+-K+ cotransport rate constants (lithium replacing sodium), intracellular sodium concentrations, and the number of Na+,K+-adenosine triphosphatase (ATPase) sites per erythrocyte. Subjects included men and women, blacks and whites. Hypertensive subjects had significantly higher sodium transport than did normotensive subjects for ouabain-sensitive sodium efflux (p less than 0.025) and Na+-Li+ countertransport (p less than 0.001). Sexual differences were noted for ouabain-sensitive (p less than 0.001) and ouabain-insensitive (p less than 0.001) sodium efflux, for intracellular sodium concentration (p less than 0.025), and for the Li+-K+ cotransport rate constant (p less than 0.005), all with higher values for men than for women. Racial differences were noted for ouabain-insensitive sodium efflux (p less than 0.005), Na+-Li+ countertransport (p less than 0.001), and the Li+-K+ cotransport rate constant (p less than 0.001); values were higher in whites than blacks for all three measurements. The number of [3H]ouabain binding sites was lower for blacks (p less than 0.001) and the intracellular sodium concentration was higher for blacks (p less than 0.001). Among all subjects, significant (p less than 0.001) correlations were found between intracellular sodium concentration and the number of Na+,K+-ATPase sites per erythrocyte (r = -0.78) and between the ouabain-sensitive sodium efflux per site and intracellular sodium concentration (r = 0.85, p less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Sodium transport parameters in erythrocytes of patients with primary aldosteronism.

Primary aldosteronism is an uncommon cause of hypertension but one of particular interest because of its distinctive pathophysiological mechanism of blood pressure elevation. Aldosterone has been associated with increased Na+,K+-adenosine triphosphatase (ATPase) activity, but there is controversy over which sodium transport parameters are responsible for this increase. We measured intracellular sodium, ouabain-sensitive and ouabain-insensitive sodium efflux, and the number of Na+,K+-ATPase sites of washed erythrocytes, as well as Na+-Li+ countertransport and the Li+-K+ cotransport rate constant of lithium-loaded red blood cells (RBCs) in six patients with primary aldosteronism and in 50 normal subjects. Ouabain-sensitive sodium efflux was significantly (p less than 0.001) higher for the primary aldosteronism patients than for normal subjects (1.85 +/- 0.29 vs 1.51 +/- 0.21 mmol/L RBC/hr) even though the intracellular sodium concentration (7.2 +/- 1.5 vs 6.7 +/- 1.9 mM) and the number of the Na+,K+-ATPase sites per RBC (331 +/- 52 vs 385 +/- 97) were not increased. The elevated sodium efflux appeared to be due to a significant (p less than 0.001) increase in the rate constant (1.60 +/- 0.12 x 10(-15) vs 1.28 +/- 0.15 x 10(-15) mmol/site/hr) of the ouabain-sensitive sodium efflux. The rate constant decreased significantly (p less than 0.01) after treatment.

Furosemide-sensitive K+ (Rb+) transport in human erythrocytes: modes of operation, dependence on extracellular and intracellular Na+, kinetics, pH dependency and the effect of cell volume and N-ethylmaleimide.

The effect of extracellular and intracellular Na+ (Nao+, Nai+) on ouabain-resistant, furosemide-sensitive (FS) Rb+ transport was studied in human erythrocytes under varying experimental conditions. The results obtained are consistent with the view that a (1 Na+ + 1 K+ + 2 Cl-) cotransport system operates in two different modes: mode i) promoting bidirectional 1:1 (Na+-K+) cotransport, and mode ii) a Nao+-independent 1:1 ki+ exchange requiring Nai+ which, however, is not extruded. The activities of the two modes of operation vary strictly in parallel to each other among erythrocytes of different donors and in cell fractions of individual donors separated according to density. Rb+ uptake through Rbo+/Ki+ exchange contributes about 25% to total Rb+ uptake in 145 mM NaCl media containing 5 mM RbCl at normal Nai+ (pH 7.4). Na+-K+ cotransport into the cells occurs largely additive to K+/K+ exchange. Inward Na+-Rb+ cotransport exhibits a substrate inhibition at high Rbo+. With increasing pH, the maximum rate of cotransport is accelerated at the expense of K+/K+ exchange (apparent pK close to pH 7.4). The apparent KmRbo+ of Na+-K+ cotransport is low (2 mM) and almost independent of pH, and high for K+/K+ exchange (10 to 15 mM), the affinity increasing with pH. The two modes are discussed in terms of a partial reaction scheme of (1 Na+ + 1 K+ + 2 Cl-) cotransport with ordered binding and debinding, exhibiting a glide symmetry (first on outside = first off inside) as proposed by McManus for duck erythrocytes (McManus, T.J., 1987, Fed. Proc., in press). N-ethylmaleimide (NEM) chemically induces a Cl--dependent K+ transport pathway that is independent of both Nao+ and Nai+. This pathway differs in many properties from the basal, Nao+-independent K+/K+ exchange active in untreated human erythrocytes at normal cell volume. Cell swelling accelerates a Nao+-independent FS K+ transport pathway which most probably is not identical to basal K+/K+ exchange. Ko+ less than Nao+ less than Lio+ less than Mgo2+ reduce furosemide-resistant Rb+ inward leakage relative to cholineo+.

FAMILIAL AGGREGATION OF RED BLOOD CELL CATION TRANSPORT SYSTEMS IN JAPANESE FAMILIES

The rate of lithium, sodium, and potassium transport in red blood cells is thought to be associated with essential hypertension. In order to investigate the contribution of genetic and environmental factors to cation transport, their correlation was analyzed in 60 parent-offspring pairs and 17 husband-wife pairs in Toyama City, Japan. Lithium-sodium countertransport and sodium-potassium cotransport rates were significantly correlated in parent-offspring pairs (r = 0.52, p less than 0.01 and r = 0.46, p less than 0.01, respectively) but not in husband-wife pairs. Sodium pump rates were significantly correlated in both pairs (r = 0.48, p less than 0.01 in parent-offspring pairs, r = 0.46, p less than 0.05 in husband-wife pairs). Therefore, lithium-sodium countertransport and sodium-potassium cotransport were found to have a substantial genetic component and the sodium pump to have a substantial environmental component. Sodium pump rates were significantly correlated with sodium/creatinine (r = 0.21, p less than 0.05) and sodium/potassium (r = 0.32, p less than 0.01) in casual urine.

Erythrocyte cation cotransport and countertransport in essential hypertension.

We studied erythrocyte cation cotransport and countertransport systems in 21 and 27 patients with essential hypertension, respectively, all of whom were under 50 years of age, had a diastolic blood pressure level greater than 100 mm Hg, and had a family history of hypertension. The following parameters were normal in nearly all patients: total erythrocyte Na+ and K+ concentrations, the maximal rate (Vmax) of inward cotransport, the affinity of cotransport with Rb+ as the substrate, the net outward cotransport of Na+ ions, the passive "leak" influx of Rb,+ and the maximal rate of Li+-Na+ countertransport. Only four patients gave clearly abnormal results; in two the maximal rate of both cotransport and countertransport was double the normal values, while another two patients demonstrated a greater than twofold increase in passive "leak" influx to Rb+ ions. Most of the patients with moderate to severe essential hypertension in this Australian study were characterized by normal erythrocyte cation fluxes, but a few showed elevation of both cotransport and countertransport of cations.

Sodium-lithium exchange and sodium-potassium cotransport in human erythrocytes. Part 2: A simple uptake test applied to normotensive and essential hypertensive individuals.

The sodium-lithium (Na+-Li+) exchange and sodium-potassium (Na+-K+) cotransport activities were assessed on erythrocytes of 38 normotensive individuals, 18 patients with well-established essential hypertension, and five renal hypertensive patients, by means of an uptake assay method. With both transport systems, no significant differences in mean values and variance were observed among the three groups. Four of six low-renin essential hypertensive patients exhibited some of the lowest exchange and cotransport rates obtained among all individuals examined. The activity of both transport systems was slightly lower in women than in men.

The involvement of the membrane oxidoreduction system in stimulating amino acid uptake in ehrlich ascites tumor cells

The addition of 5 mM ascorbate plus 0.09 mM phenazine methosulfate stimulated 2− to 3−fold the initial rate of 2-aminoisobutyric acid transport into Ehrlich cells. This was observed under the conditions in which glycolysis and mitochondrial electron transport were blocked by iodoacetate and KCN, and the cellular ATP level was maintained below 0.1 mM. Proton conductors, carbonylcyanide m- chlorophenylhydrazone and SF6847 did not affect the stimulation of 2-aminoisobutyric acid uptake caused by ascorbate plus phenazine methosulfate. Ascorbate was replaced by NADH but not by NADPH, and phenazine methosulfate was the only effective acceptor in stimulating 2-aminoisobutyric acid uptake. The stimulating effect of ascorbate plus phenazine methosulfate was due to an increase in the V value for 2-aminoisobutyric acid but not in the K m value. This effect required the presence of an Na + gradient and was accompanied by an increase in 22Na + influx. The molar ratio of 2-aminoisobutyric acid to Na + uptake enhanced by ascorbate plus phenazine methosulfate was calculated to be 1 : 1. Quinacrine, an inhibitor of NADH oxidoreductase in the plasma membrane, inhibited both the enhanced rate of 2-aminoisobutyric acid and Na + transport without affecting the basal transport activity. The stimulatory effect of ascorbate plus phenazine methosulfate was also observed with other amino acids, alanine, glycine, proline and cycloleucine which are known to be transported via an Na + dependent system but not with leucine and threonine. These results suggest that a redox system in the plasma membrane participates in energy coupling for amino acid transport by increasing the rate of cotransport with Na +.