Ouabain-sensitive Efflux Rate Constant Research Articles

Alteration in ouabain-sensitive sodium potassium pump of erythrocytes during pregnancy induced hypertension: a kinetic study.

The erythrocytes are widely used as model cells for studies of sodium-potassium pump (Na(+)-K(+) pump) in health and disease. Hence, to explore the possible role of the Na(+) transport across the cell membrane in the pathogenesis of pregnancy-induced hypertension (PIH), the present study was conducted to assess the Na(+)-K(+) pump functions in relation to its intrinsic kinetic properties using erythrocytes (RBC). Erythrocyte sodium concentration in pregnancy-induced hypertensive women was significantly (p<0.01) lower in comparison to normotensive pregnant women. On the contrary erythrocyte potassium was significantly higher (p<0.01) in PIH women as compared to normotensive pregnant women. Observed alterations in Na(+) and K(+) concentrations in erythrocytes were associated with significantly (p<001) increased Ouabain-sensitive sodium efflux rate and rate constants in erythrocytes from PIH women. Further, kinetic studies revealed that increased Ouabain-sensitive efflux rate constant in RBC from PIH women was accompanied by increased maximal velocity (V(max)) of Na(+)-K(+) pump. However, the affinity constant (K(m)) was unaltered in both the groups. Therefore, these findings suggest that increased Na(+)-K(+) pump activity in RBC of PIH women could be due to either increased numbers of Na(+)-K(+) pump units of increased numbers of active subunits of Na(+)-K(+) pump possibly due to specific plasma factors in PIH women.

Leucocyte sodium efflux and electrolyte content in insulin treated diabetes.

Leucocyte sodium efflux and sodium content were studied in 41 insulin treated diabetic patients and compared to 41 age, body mass index and blood pressure matched nondiabetic control subjects. Fasting leucocyte 22Na ouabain-sensitive efflux rate constants were lower in diabetic patients (median [range] 2.30 [1.04-3.73] versus 2.45 [1.57-3.95] h-1, p less than 0.4) suggesting a reduced sodium pump activity. The 22Na ouabain-insensitive efflux rate constant which reflects passive sodium efflux was raised in insulin treated diabetes (0.92 [0.42-1.73] versus 0.79 [0.28-1.49] h-1, p less than 0.01). Leucocyte sodium content was raised in the diabetic patients (47.7 [26.9-93.4] versus 26.5 [15.9-67.7] mmol/kg, p less than 0.0001). Abnormal cellular sodium handling could lead to hypertension or other complications in diabetes.

Leucocyte sodium pump activity after meals or insulin in normal and obese subjects: cause for increased energetic efficiency in obesity?

As cellular sodium pumping is an energy consuming process and differences in the obese may account for their energetic efficiency, leucocyte sodium-22 efflux was studied in obese and normal volunteers both in the fasting state and after a test meal or infusion of glucose and insulin intravenously. The 22Na ouabain sensitive efflux rate constant was significantly higher in obese subjects than normal (mean (1 SD) 2.69 (0.40)/h v 2.35 (0.49)/h). Two hours after a 4.2 MJ (1000 kcal) meal there was an increase in the efflux rate constant from its fasting value in normal weight subjects (2.39 (0.33)/h to 2.71 (0.40)/h) but not in obese subjects (2.65 (0.54)/h to 2.61 (0.58)/h). The rise in ouabain sensitive efflux rates was significantly higher in normal than obese subjects. Both groups showed a rise in intracellular sodium concentrations. The euglycaemic clamp produced similar results. Feeding or infusion of insulin increases sodium pump activity more in normal than obese subjects. This difference may contribute to any defective dietary thermogenesis in obesity, which may lead to energetic efficiency and a tendency to gain weight.

THE EFFECT OF ORAL GLUCOSE ON THE LEUCOCYTE SODIUM PUMP IN NORMAL AND OBESE SUBJECTS

The effect of oral glucose (40 g/m2 body surface area) on the leucocyte 22Na efflux rate constants (ERC) was studied in 13 normal weight and 10 obese subjects. The ouabain-sensitive 22Na ERC was higher in leucocytes isolated from fasting obese subjects (median [range] for obese 2.77 [2.33-3.11] vs normals 1.91 [1.57-2.77] h-1, P less than 0.001). There was no difference in the ouabain-resistant 22Na ERC. Oral glucose raised the ouabain-sensitive 22Na ERC after 2 h in normal subjects (1.91 [1.57-2.77] to 2.41 [2.11-3.02] h-1, P less than 0.001). The ouabain-resistant 22Na ERC fell from 0.71 [0.32-1.10] to 0.46 [0.35-0.68] h-1, P less than 0.008. Conversely, in obese subjects, the ouabain-sensitive ERC fell (2.77 [2.33-3.11] to 2.59 [2.11-2.92] h-1, P less than 0.06). There was no significant change in ouabain-resistant 22Na ERC 2 h after oral glucose. The fasting leucocyte 22Na ouabain-sensitive ERC correlated with fasting plasma insulin levels and insulin resistance (rs = 0.48, P less than 0.01 for both). The change in this ERC with oral glucose correlated with the incremental insulin response over 2 h (rs = -0.53, P less than 0.006) and to the insulin resistance (rs = -0.56, P less than 0.003). The failure of oral glucose to stimulate the leucocyte sodium pump in obesity could partially account for the defect in dietary thermogenesis in obesity. This defect in stimulation of the sodium pump is related to insulin resistance.

Volume, sodium content and sodium efflux of human mononucleated cells: Characteristics and methodological problems

New methods for determining the volume and sodium content of human mononucleated cells are presented and the kinetics of sodium efflux are characterized. Sodium efflux obeyed first-order kinetics with a half-life of about 9 min. The ouabain-sensitive sodium pump is responsible for about 60% of the total turnover of this cation. Reliable measurement of cell volume and sodium content requires pre-incubation at physiological conditions. Determination of sodium efflux should be based on the initial (less than 15 min) cellular delay of 22Na-activity in preloaded cells. In normal subjects (n = 34), mean cell volume was 635 +/- 138 nl/10(6) cells and sodium content averaged 14.6 +/- 2.8 mmol/l cell. Total and ouabain-sensitive efflux rate constants were 4.56 +/- 0.66 h-1 and 2.70 +/- 0.47 h-1. Corresponding absolute sodium efflux averaged 65.1 +/- 14.8 and 38.9 +/- 9.6 mmol/l/h, respectively.

Volume, sodium content and sodium efflux of human mononucleated cells: Characteristics and methodological problems

New methods for determining the volume and sodium content of human mononucleated cells are presented and the kinetics of sodium efflux are characterized. Sodium efflux obeyed first-order kinetics with a half-life of about 9 min. The ouabain-sensitive sodium pump is responsible for about 60% of the total turnover of this cation. Reliable measurement of cell volume and sodium content requires pre-incubation at physiological conditions. Determination of sodium efflux should be based on the initial (less than 15 min) cellular delay of 22Na-activity in preloaded cells. In normal subjects (n = 34), mean cell volume was 635 +/- 138 nl/10(6) cells and sodium content averaged 14.6 +/- 2.8 mmol/l cell. Total and ouabain-sensitive efflux rate constants were 4.56 +/- 0.66 h-1 and 2.70 +/- 0.47 h-1. Corresponding absolute sodium efflux averaged 65.1 +/- 14.8 and 38.9 +/- 9.6 mmol/l/h, respectively.

The effect of dietary polyunsaturated fat on cation transport and hypertension in the rat.

1. Essential hypertension is associated with increased intracellular sodium in both erythrocytes and leucocytes. Reports in the literature indicate that increasing the level of polyunsaturated fat in the diet reduces hypertension. In the present study, spontaneously hypertensive rats (Wistar-Kyoto, which develop systolic blood pressures in excess of 140 mmHg by 8 weeks of age) were fed on high-fat diets (40% energy derived from fat), the fat being maize oil (high in polyunsaturated fatty acids, PUFA) and coconut oil (low in PUFA). 2. Significantly higher blood pressures developed by 110 d of age in the rats fed on a high-PUFA diet, compared with those fed on the low-PUFA diet. 3. In thymocytes, ouabain-sensitive efflux rate constants were significantly lower in the group fed on the high-PUFA diet. Ouabain-insensitive efflux rate constants were unaffected by diet.

Erythrocyte sodium content, sodium transport, ouabain binding capacity and Na+, K+-ATPase activity in lean and obese subjects.

Erythrocyte sodium content, sodium transport (ouabain-sensitive efflux rate of sodium, and ouabain-sensitive efflux rate constant of sodium) 3H ouabain binding capacity and sodium-potassium-activated ouabain-sensitive adenosine triphosphatase (Na+-K+-ATPase) activity were measured in 18 lean subjects and 25 obese subjects. The mean erythrocyte sodium content, sodium transport and ouabain binding capacity of obese subjects were the same as in lean subjects. There was no relationship between obesity index (wt/ht2) and sodium transport. We conclude that erythrocyte sodium transport in most obese patients is normal.

Characteristics of sodium transport in pig ( Sus scrofa) erythrocytes

Sodium content, sodium transport (ouabain-sensitive efflux rate of sodium, o M Na os; and ouabain-sensitive efflux rate constant of sodium, o K Na os), [ 3H]ouabain binding capacity, and Na +, K + -ATPase activity were measured in erythrocytes from young pigs ( Sus scrofa). The sodium content, sodium transport, and the number of sodium pumps (assessed by ouabain binding capacity) were lower in the pig compared to the human erythrocytes. The efflux rate constant of sodium, o K Na os was 36% and the ouabain binding capacity was 60% of those in human, suggesting the degree of activation of sodium pump units is much lower.

Effect of hypokalaemia on the ouabain-sensitive sodium transport and the ouabain-binding capacity in human erythrocytes.

1. The sodium content, the ouabain-sensitive sodium efflux and efflux rate constant and the ouabain-binding capacity were measured in the erythrocytes of 53 patients with hypokalaemia and in 37 healthy controls. The sodium content alone was measured in a further 57 patients with hypokalaemia. 2. In the patients with hypokalaemia there was an increase in the average sodium content of the erythrocytes, which was entirely due to a reduction in the ouabain-sensitive efflux rate constant. 3. The ratio of the ouabain-sensitive efflux rate constant to the number of sodium pumps was decreased in the patients with hypokalaemia, and was directly related to the plasma potassium. 4. Many patients with moderate hypokalaemia had normal erythrocyte sodium and potassium contents and normal ouabain-sensitive efflux rate constant. These patients had an increased number of sodium pumps, which compensated for the inhibitory effect of hypokalaemia on each sodium pump. 5. This increase in the number of sodium pumps was common even in patients who had probably had hypokalaemia for less than 2 weeks. This finding suggests that there are latent sodium pumps within the circulating erythrocytes.

The Effect of Pregnancy and Pregnancy-Induced Hypertension on Active Sodium Transport in the Erythrocyte

Abstract 1. (1) Active sodium transport was assessed in the erythrocytes of pregnant women with and without pregnancy-induced hypertension and in non-pregnant women. 2. (2) The measurements made were the sodium content of washed erythrocytes (Naw), the ouabain-sensitive efflux rate constant (Kos), the ouabain-sensitive sodium flux (Mos) and the number of sodium pumps (as the ouabain-binding capacity; OB). 3. (3) The sodium content was reduced in the pregnant women due to an increase in the efflux rate constant and this increase was due to an increased number of sodium pumps (by 50%) which swamped a 20% reduction in the rate constant per pump. 4. (4) There was a general reduction in the permeability of the erythrocyte to sodium but at the same time there was an increase in permeability in association with the increased number of sodium pumps. The greater variability of permeability and therefore of sodium flux in the pregnant women was explained by the variable increase in the number of sodium pumps. 5. (5) There were no significant differences in these measurements between the hypertensive and the normotensive pregnant women and in particular there was no difference in the degree of pump inhibition. 6. (6) These results suggest that the major change in active sodium transport in pregnancy is an increase in the number of sodium pumps. We suggest that this is a part of the wide ranging effect of pregnancy on protein synthesis and not a specific effect on sodium transport. There is an inhibition of the sodium pump in pregnancy but it does not cause an increase in the sodium content of the erythrocytes because there is also an increase in the number of sodium pumps. The inhibition is no greater in hypertensive than in normotensive pregnancies.

Erythrocyte sodium transport in chronic renal failure.

1. Erythrocyte sodium, sodium transport (ouabain-sensitive efflux rate of sodium, oMosNa, and ouabain-sensitive efflux rate constant of sodium, oMosNa), sodium-potassium activated ouabain-sensitive adenosine triphosphatase (Na+, K+-ATPase) activity and [3H]ouabain-binding capacity were measured in 15 patients with chronic renal failure and in 10 healthy subjects. 2. As a group, patients with chronic renal failure had a lower erythrocyte sodium and oMosNa compared with healthy subjects. 3. When patients were divided according to their erythrocyte sodium (greater or less than 4 mmol/kg of cells), in the group of patients whose erythrocyte sodium was less than 4 mmol/kg of cells (group A) the oMosNa was higher than that in healthy subjects and the oMosNa, Na+, K+-ATPase activity and [3H]ouabain-binding capacity were the same as those in healthy subjects. In the subgroup of patients with renal failure whose erythrocyte sodium content was greater than 4 mmol/kg of cells (group B) the oMosNa was less and plasma urea concentration higher than in group A and Na+, K+-ATPase activity, [3H]ouabain-binding capacity and oMosNa were lower than in healthy subjects. 4. These results suggest that in early chronic renal failure there is stimulation of ‘sodium pumps’ (without alteration in their number), which causes a lowering of erythrocyte sodium content, and that as the disease progresses there is inhibition of the ‘sodium pumps’ as well as a reduction in membrane permeability so that erythrocyte sodium is near normal.

The relationship between sodium transport and Na +, K + ATPase in human erythrocytes

Erythrocyte sodium, potassium-activated, ouabain-sensitive adenosine triphosphatase (Na +, K + ATPase) and the ouabain-sensitive efflux rate constant of sodium (ERC os) and sodium content ([Na] RBC ) were measured in six groups of subjects. There were 25 healthy young adults, 15 elderly subjects, 16 patients with hyperthyroidism, 6 patients during digitalization, 8 patients who had been on digoxin for more than 14 months, and 25 patients with chronic renal failure. In the healthy subjects there was a significant correlation between the Na +, K + ATPase activity and the ERC os ( r = 0.589) and between [Na] RBC and Na +, K + ATPase ( r = −0.695). There was also a highly significant correlation ( r = 0.91) between the mean values of ERC os and Na +, K + ATPase for the six groups. There results add further confirmation of the close relationship between the activity of the enzyme and sodium transport.