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L M Sakhrani

Publications and source records attributed to L M Sakhrani.

12 recordsLinked to original sources
PubMed1986

Quantitative requirement for ATP for active transport in isolated renal cells.

We investigated the quantitative relationship between cellular ATP concentration and Na+-K+-ATPase activity as measured by ouabain-sensitive 86Rb influx in rabbit proximal renal cells. Cellular ATP was reduced in a stepwise manner by rotenone (10(-7) to 10(-5) M) and was increased by 10 mM adenosine. During these maneuvers, ouabain-sensitive 86Rb influx was linearly related to cellular ATP and did not saturate up to 9.9 mM ATP. In contrast, Na+-K+-ATPase activity in membranes prepared from these cells saturated at 2.0 mM ATP at various sodium (10-100 mM) and potassium (4-100 mM) concentrations. Sodium-dependent phosphate uptake and alpha-methylglucoside (alpha-MG) uptake were both inhibited to a similar degree when cellular ATP was reduced. We conclude that 1) the ATP requirement for saturation of Na+-K+-ATPase is higher in intact renal cells than in the membranes, and 2) the uptake of phosphate and alpha-MG are similarly influenced by reduction in ATP. This effect of ATP on phosphate and AMG uptake is most likely an indirect one and is secondary to changes in the sodium gradient across the cell.

Adenosine Triphosphate↗
PubMed1986

Proximal tubular cells in primary culture.

Primary cultures of mammalian proximal tubules (PT) can be established in serum-free, hormone-supplemented media. These cells attain confluence, demonstrate morphological polarity and exhibit vectorial fluid transport. Primary cultures of rabbit PT cells retain the transport properties of the parent cells and exhibit Na+-H+ exchange, Na+-Ca2+ exchange and Na-glucose co-transport, the latter process having kinetic properties comparable to those described for the late proximal tubule. Glucose which enters the cell via the luminal Na-glucose co-transport system undergoes metabolism to CO2; it is not clear whether this glucose oxidation, which is minimal in PT cells under physiological conditions, is due to selection of a specific cell type or to the nature of the culture conditions. Confluent, quiescent PT cells in culture can be induced to hypertrophy by addition of supraphysiological concentrations of insulin, PGE, and hypertonic NaCl; each stimulus also leads to an early increase in Na+-H+ antiport suggesting that this transport process may be causally related to the process of cell growth. The primary culture thus represents a promising model for future studies on the cellular events which govern the process of cell hypertrophy.

Adenylyl Cyclases↗
PubMed1986

Effect of dopamine on sodium uptake by renal proximal tubule cells of rabbit.

Although dopamine is known to be natriuretic, it is not clear if this is due to changes in renal hemodynamics or to a direct tubular effect. Studies on the effect of dopamine on proximal sodium reabsorption have yielded conflicting information, both an increase and a decrease in sodium reabsorption has been reported. The present study examines the direct effect of dopamine on sodium uptake in proximal renal cells and the possible mechanisms involved. Dopamine (10(-7)-10(-4) M) stimulated in a dose-dependent manner sodium uptake by proximal renal cells by 35-92%; 1 mM ouabain and 70 microM cycloheximide did not modify the effect of dopamine. Kinetic analysis of sodium uptake by these cells showed a single saturable component, inhibitable by amiloride, with a Km of 80 +/- 6 mM and Vmax of 68 +/- 9 pmol/mg protein/min. Dopamine (10(-4) M) increased the Vmax of sodium uptake by 54 +/- 10.3% and had no effect on the Km. Metoclopramide abolished the stimulatory effect of dopamine on sodium uptake whereas propranolol had no effect. Epinine (a dopamine agonist) also stimulated sodium uptake by these cells. We conclude that dopamine directly stimulates sodium uptake in proximal renal cells; this suggests the natriuretic effect of dopamine in whole animals is due to changes in renal hemodynamics and distal tubular effects; dopamine increases the Vmax of the sodium transporter and not the Km; the sensitivity of sodium uptake to amiloride suggests that dopamine stimulates Na+-H+ exchanger, and the stimulation of sodium uptake by dopamine is independent of Na+-K+-ATPase and new protein synthesis and occurs via the dopamine receptor.

Animals↗
PubMed1985

Effect of calcium on transport characteristics of cultured proximal renal cells.

We examined the effects of acute changes in extracellular and intracellular calcium on transport processes in primary culture of proximal rabbit renal cells. A change in extracellular calcium from 0 to 3 mM inhibited amiloride-sensitive sodium uptake by 30%, and this effect was maximal at 1 mM calcium. Other polyvalent cations (Mn2+, Mg2+, La3+, and Ba2+) produced quantitatively similar inhibition of amiloride-sensitive sodium uptake compared with calcium. An increase in cytosolic calcium produced by calcium loading (20 mM) or by A23187 (20 microM) resulted in an inhibition of 25-40% of amiloride-sensitive sodium uptake. Moreover, quinidine (10(-4)M) and ruthenium red (3 microM), agents presumed to increase cytosolic calcium, inhibited amiloride-sensitive sodium uptake by 20-60%. Both these agents also inhibited sodium-dependent phosphate uptake by 20% but had no effect on ouabain-sensitive 86Rb+ uptake or on sodium-dependent alpha-methylglucoside uptake. Our data indicate that increases in extracellular calcium inhibit amiloride-sensitive sodium uptake and increases in cytosolic calcium inhibit sodium-dependent phosphate and amiloride-sensitive sodium uptakes. The effect of extracellular calcium may be due to charge screening and/or binding to the negatively charged plasma membrane or due to alterations in membrane fluidity.

Amiloride↗
PubMed1985

Hyponatremia.

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Blood Volume↗
PubMed1985

Interactions between tricarboxylic acid cycle intermediates and phosphate uptake by proximal renal cells and renal brush border membranes.

Glucose and other hexoses as well as amino acids have been shown to inhibit the renal transport of phosphate (Pi). Although studies with renal brush border membrane vesicles showed that such an inhibitory effect on Pi transport is due to the dissipation of the Na electrochemical gradient, the mechanism(s) responsible for such an action in the intact cell is not clear. The present study examined the effects of tricarboxylic acid (TCA) cycle intermediates (succinate and fumarate) and acetate on the uptake of Pi and alpha-methylglucoside (AMG) at 37 degrees C by intact rabbit renal cells. These TCA cycle compounds significantly (p less than 0.05) inhibited the uptake of both Pi and AMG. In the presence of 5-10 mM succinate the ATP content of the renal cells increased by 40% (p less than 0.02). Inhibition of succinate-induced gluconeogenesis by 3-mercaptopicolinic acid did not modify the inhibition of Pi uptake. Studies with renal brush border membrane vesicles showed that succinate inhibited Pi uptake at 15 and 60 s but not at 1 s and only under conditions of Na gradient (outside greater than inside). Succinate did not inhibit Pi uptake during Na equilibrium conditions. The data demonstrate that the succinate-induced inhibition of the Pi uptake by intact proximal renal cells is not due to competition for metabolic energy, is not related to stimulation of gluconeogenesis nor due to allosteric interaction between Pi carrier and succinate transporter. The results support the notion that the inhibition of Pi uptake by succinate in the intact renal cell is due to dissipation of the Na chemical gradient.

Animals↗
PubMed1984

Transport and metabolism of glucose by renal proximal tubular cells in primary culture.

A highly purified suspension of rabbit proximal tubules was cultured in a hormone-supplemented serum-free medium. This primary culture yielded a homogeneous population of cells that demonstrated functional and morphological polarity in mono-layers. The characteristics of the Na-dependent glucose transporter in the luminal membrane were studied by measuring the uptake of alpha-methylglucoside (AMG). The kinetics of Na-dependent AMG uptake were consistent with a single saturable system with an apparent Km of 0.8 mM and Jmax of 0.14 nmol X mg-1 X min-1. AMG permeability was 0.10 microliter X mg-1 X min-1. Uptake was inhibited 95% by 0.1 mM phlorizin and by removal of sodium. The stoichiometry of Na/glucose interaction with the carrier was 2:1. These characteristics are typical of the characteristics described for the late proximal tubule. To examine whether the glucose that enters the cell across the luminal membrane is incorporated into the metabolic pool of the cell, we studied the oxidation of [14C]glucose to 14CO2 in the absence and presence of phlorizin. Significant decarboxylation of [1-14C]glucose and [6-14C]glucose was observed, consistent with the existence of aerobic metabolism and a hexose monophosphate shunt. In the presence of 0.1 mM phlorizin, uptake and oxidation of D-glucose were inhibited to an identical degree, suggesting that luminal uptake is a rate-limiting step in the oxidation of glucose by these proximal tubular cells. These studies indicate that proximal tubular cells in primary culture utilize glucose as an energy source and that the glucose derived from transport across the luminal membrane is incorporated into the metabolic pool of the cell.

Animals↗
PubMed1984

Role of substrates and nucleotides in phosphate uptake by rabbit renal cortical cells.

We investigated the viability of isolated rabbit renal cortical cells and the effect of metabolic intermediates on phosphate uptake. Glucose production and its stimulation by hormones was similar to that previously reported for renal tubules. The uptake of alpha-methylglucoside and phosphate was reduced by 90% when sodium was removed from the media. ATP content of the cells was 9.4 +/- 0.7 nmol/mg protein. Succinate and fumarate stimulated phosphate uptake in a dose-dependent manner by 30-60%. Valeric acid (1 mM) and butyric acid (10 mM) stimulated phosphate uptake by 20-30%. Glucose and fructose stimulated phosphate uptake by 18% but only at low concentrations (0.1 mM). Exogenous nucleotides had no effect on phosphate uptake at 0.2 mM, but inhibited 2.4 mM phosphate uptake at 2 mM. We conclude: rabbit renal cortical cells in suspension are metabolically and functionally viable; tricarboxylic acid cycle and glycolytic intermediates as well as short chain fatty acids can stimulate phosphate uptake, and exogenous nucleotides are hydrolyzed to produce free phosphate causing inhibition of isotopic phosphate uptake.

Animals↗