Peritubular Cell Membrane Research Articles

Electrophysiological analysis of bicarbonate permeation across the peritubular cell membrane of rat kidney proximal tubule. II. Exclusion of HCO3(-)-effects on other ion permeabilities and of coupled electroneutral HCO3(-)-transport.

Cell membrane potentials of rat kidney proximal tubules were measured in response to peritubular ion substitutions in vivo with conventional and Cl- sensitive microelectrodes in order to test possible alternative explanations of the bicarbonate dependent cell potential transients reported in the preceding paper. Significant direct effects of bicarbonate on peritubular K+, Na+, and Cl- conductances could be largely excluded by blocking K+ permeability with Ba2+ and replacing all Na+ and Cl- by choline or respectively SO4(2-) isethionate, or gluconate. Under those conditions the cell membrane response to HCO3- was essentially preserved. In addition it was observed that peritubular Cl- conductance is negligibly small, that Cl-/HCO3- exchange - if present at all - is insignificant, and that rheogenic HCO3- flow with coupling to Na+ flow is also absent or insignificant. A transient disturbance of the Na+ pump or a transient unspecific increase of the membrane permeability was also excluded by experiments with ouabain and by the observation that SITS (4-acetamido-4'-isothiocyano-2,2' disulphonic stilbene) blocked the HCO3- response instantaneously. The data strongly support the notion that the potential changes in response to peritubular HCO3- concentration changes arise from passive rheogenic bicarbonate transfer across the peritubular cell membrane, and hence that this membrane has a high conductance for bicarbonate buffer.

Evidence for an amiloride sensitive Na+ pathway in the amphibian diluting segment induced by K+ adaptation.

The effect of amiloride on cell membrane potentials and intracellular Na activity (Nai) was tested in early distal tubules of the isolated perfused kidney of control and of K-adapted (high-K diet) Amphiuma. Conventional and Na-sensitive liquid ion-exchanger microelectrodes were employed to measure the peritubular cell membrane potential (PDpt), the transepithelial potential difference (PDte) and the Na electrochemical gradient across the peritubular cell membrane (ENapt), in the absence and the presence of amiloride (1 X 10(-4) mol X 1(-1] in both groups of animals. Amiloride did not affect PDpt and ENapt in control animals but depolarized PDpt and ENapt by about 8 mV in K-adapted animals. Nai (11.0 +/- 0.6 mmol X 1(-1) in early distal cells of control animals) did not change significantly by this maneuver. However, Nai decreased to extremely low values (2.3 +/- 0.2 mmol X 1(-1] when the luminal cotransport system for Na, Cl and K was inhibited by the luminal application of furosemide (5 X 10(-5) mol/l) and when the luminal cell membrane was exposed simultaneously to amiloride. The amiloride-induced effects on PDpt, ENapt and Nai occurred within seconds and were fully reversible. We conclude that high-K diet (K adaptation) induces an amiloride-sensitive pathway in the luminal cell membrane of early distal cells of Amphiuma which exists in parallel with the furosemide-sensitive cotransport system located in this cell barrier. The results suggest a luminal amiloride-sensitive Na/H exchange mechanism which regulates the luminal K permeability.

Interaction of intracellular electrolytes and tubular transport.

To disclose possible regulatory mechanisms, the potential difference across the peritubular cell membrane (PDpt) and intracellular activities of sodium (Nai+), potassium (Ki+), calcium (Cai2+), bicarbonate (HCO3i-) and chloride (Cli-) have been traced continuously during inhibition of Na+/K+-ATPase with ouabain. Within 31 +/- 4 min following application of ouabain, PDpt decreases (from 57 +/- 2 mV) to half and Ki+ by 37.7 +/- 2.2 mmol/l (from 63.5 +/- 1.9 mmol/l), Nai+ increases by 35.1 +/- 4.1 mmol/l (from 13.2 +/- 2.4 mmol/l), Cai2+ by 0.17 +/- 0.2 mumol/l (from 0.09 mumol/l), HCO3i-) by 3.0 +/- 1.1 mmol/l (from 15.3 +/- 2.0 mmol/l) and Cli- by 6.2 +/- 1.0 mmol/l (from 14.4 +/- 1.6 mmol/l). Within the same time the luminal and peritubular cell membrane resistances increase 45 +/- 15% and 53 +/- 17%, respectively. The increase of the resistances is mainly due to a decrease of K+ conductance, which in turn mainly accounts for the depolarisation of PDpt. Additional experiments demonstrate that the K+ conductance of the peritubular cell membrane is sensitive to the cell membrane potential difference and possibly linked to Na+/K+-ATPase activity. The decline of PDpt probably accounts for intracellular alkalinisation which in turn reduces Na+/H+ exchange. Na+-coupled transport of glucose and phenylalanine decrease in linear proportion to PDpt. The transport of these and probably of similar substances represents the main threat to electrolyte homeostasis of the cells.

Anthracene-9-carboxylic acid inhibits renal chloride reabsorption.

From previous studies, it is known that in the diluting segment, C1- -ions are transported from the tubule lumen into the cell together with Na+ and K+ via a furosemide-sensitive cotransport system. This carrier-mediated process, located in the luminal cell membrane, is driven by the steep "downhill" Na+ gradient (directed from lumen to cell) which is maintained by the ouabain-sensitive Na+/K+-pump at the peritubular cell membrane. C1- -ions are accumulated within the cell cytosol and are supposed to leave the cell by a C1- -conductive pathway. The present experiments, performed in diluting segments of the isolated perfused frog kidney, demonstrate the existence of a significant C1- -permeability of the peritubular cell membrane and its complete inhibition by anthracene-9-COOH. The data indicate that C1- -reabsorption can be reduced not only by the inhibition of luminal C1- -entry (i.e. by furosemide) but also by the blockade of the passive C1- -exit step across the peritubular cell membrane. Since complete inhibition of C1- -permeability reduces transepithelial uphill C1- -transport only to half, the data disclose the existence of an additional C1- -pathway at the peritubular cell membrane.

Effect of luminal potassium on cellular sodium activity in the early distal tubule of Amphiuma kidney.

From previous studies it is known that a furosemide-sensitive sodium chloride cotransport system is operative in the luminal cell membrane of the early distal amphibian tubule. Since inhibition of sodium chloride cotransport prevents potassium reabsorption in this nephron segment, experiments were carried out to evaluate further the possible relationship between sodium chloride and potassium transport by studying the changes of cellular sodium activity following luminal deletion of potassium ions. Sodium-sensitive liquid ion exchange microelectrodes and conventional microelectrodes were employed to determine the transepithelial potential (PDte), the peritubular cell membrane potential (PDpt) and the intracellular sodium activity (Nai+) in the presence and absence of luminal potassium. The ratio of the luminal cell membrane resistance over the peritubular cell membrane resistance (Rlu/Rpt) was also estimated. When potassium ions are omitted from the luminal perfusate, PDpt hyperpolarizes by some 20 mV, PDte approaches zero and Nai+ decreases by about 40%. Rlu/Rpt is more than doubled in the presence of a potassium-free perfusate. Both potential and resistance changes are fully reversible. Similar results were obtained in experiments in which Barium ions (1 mmol/1 BaCl2) were present during the luminal potassium substitution. Our results indicate that absence of potassium inhibits luminal sodium chloride entry; as a result of continued peritubular sodium extrusion cellular sodium activity falls. The increase of Rlu/Rpt following perfusion with a potassium-free perfusate is interpreted as a decrease of a significant electrodiffusive potassium conductance in the luminal cell membrane.

The effect of cAMP on the cell membrane potential and intracellular ion activities in proximal tubule of Rana esculenta.

Experiments were performed in proximal tubule of the isolated perfused frog kidney to evaluate peritubular cell membrane potentials (PDpt), and the intracellular ion activities of sodium (Nai+), chloride (Cli-) and potassium (Ki+) under control conditions and following peritubular application of dibutyryl-cyclic AMP (cAMP, 2 X 10(-4) mol X 1(-1)). Conventional and ion-sensitive microelectrodes were applied to record continuously cAMP-induced changes of these parameters in individual proximal tubule cells. Within a few minutes a significant hyperpolarisation of PDpt (delta = 2.0 +/- 0.2 mV) occurs simultaneously with a decrease of Nai+ (delta = 2.5 +/- 0.5 mmol X 1(-1)). Ki+ increases (delta = 3.6 +/- 0.9 mmol X 1(-1)) and Cli- decreases (0.4 +/- 0.07 mmol X 1(-1)) slightly, but significantly. With both ions the alterations of the chemical gradient is significantly smaller than the potential shift. PDte is not significantly altered by cAMP. The cAMP-induced hyperpolarisation of PDpt can be observed in presence and absence of luminal glucose. However, omission of Na+ from the luminal perfusate abolishes the hyperpolarising effect of cAMP on PDpt. The results suggest that cAMP reduces sodium entry from the lumen into the cell, thus hyperpolarising the cell membrane and decreasing Nai+. Persistence of sensitivity of PDpt to cAMP after omission of glucose indicates that other Na+ coupled transport processes and/or passive Na+ conductance are affected by cAMP. the changes of Ki+ and Cli- are secondary, following the change of PDpt.

Omission of luminal potassium reduces cellular chloride in early distal tubule of amphibian kidney.

Experiments in the amphibian early distal tubule have shown that Cl transport is secondarily active, coupled to the flux of Na and dependent on the presence of luminal K. Omission of luminal K results in a decrease of cellular Na, a finding that suggests inhibition of luminal Na entry. In the present study intracellular chloride activity (Cli) and peritubular cell membrane potentials (PDpt) were evaluated before and after omission of luminal K. Furthermore, the effect of inhibition of the luminal K conductance by barium on the electrochemical gradient of Cl (E te Cl ) across the distal epithelium was determined at static head conditions. Experiments were performed in early distal segments of the isolated perfused kidney ofAmphiuma andRana esculenta. Cli and PDpt were measured simultaneously in single cells by double barreled Cl sensitive microelectrodes in the presence and absence of luminal K. E te Cl was determined at zero net flux conditions with single barreled electrodes in control tubules, in the presence of barium (3·10−3 mol/l) and in the presence of furosemide (5·10−5 mol/l). In 26 individual cellular impalements omission of luminal K hyperpolarized PDpt from 72.5±1.2 to 90.0±1.9 mV (cell interior negative). Concomitantly, Cli fell from 8.5±0.4 to 5.4±0.3 mmol/l. Both effects occurred within seconds and were fully reversible. Addition of barium to the luminal fluid diminished E te Cl (directed lumen positive) from a control value of 39.5±1.4 mV to 28.5±2.5mV. E te Cl could be further diminished to 14.1±2.1 mV and to 1.3±0.5 mV after application of barium on both sides and after luminal application of furosemide, respectively. The experiments indicate that active Cl uptake across the luminal cell membrane depends critically on the presence of luminal K. Omission of luminal K achieved either by perfusing the lumen with K-free solutions or by inhibition of K back flux from the cell interior into the lumen by barium reduces Cl reabsorption. Together with previous data on the K dependence of the Na uptake the present experiments support the hypothesis of a common transport system for K, Na, Cl located in the luminal cell membrane.

Cellular Mechanism of the furosemide sensitive transport system in the kidney.

Experiments were performed in the distal tubule of the doubly-perfused kidney of Amphiuma to determine active and passive forces, involved in the transport processes of potassium, sodium and chloride. Ion-sensitive microelectrodes and conventional microelectrodes were applied to estimate intracellular ion activities, cell membrane potentials and net flux of potassium and chloride under control conditions and during inhibition of active transport. Sodium chloride cotransport, located in the luminal cell membrane is postulated, based on the following observations: Total omission of sodium from the tubular lumen inhibits furosemide sensitive chloride reabsorption, decreases the lumen positive transepithelial potential difference and leads to a dramatic decrease of intracellular chloride. The experiments further suggest that potassium ions are involved in the sodium chloride transport system because potassium reabsorption is inhibited by furosemide and because intracellular sodium falls significantly when potassium ions are removed from the tubular fluid. Furthermore, there is experimental evidence that the luminal potassium uptake mechanism is suppressed after potassium adaptation. Under these conditions potassium transport is found to be insensitive to furosemide. The data suggest a furosemide sensitive cotransport system for sodium, chloride and potassium, operative in the luminal cell membrane. The energy for this carrier-mediated transport process is provided by the large "downhill" gradient of sodium across the luminal cell membrane which is maintained by the sodium pump located in the peritubular cell membrane.

Electrophysiological analysis of rat renal sugar and amino acid transport. V. Acidic amino acids.

We have used electrophysiological techniques to study various aspects of the transport of glutamate and aspartate in proximal tubules of the rat kidney in vivo. Single tubular cells were punctured with microelectrodes and the response of the cell membrane potential to sudden luminal or peritubular applications of these amino acids was measured. The experiments indicated that a specific transport system exists for L-glutamate and L-aspartate in the brushborder membrane, which does not transport neutral or basic amino acids. The uptake of both L-amino acids from the lumen into the cell was found to be rheogenic, probably reflecting the cotransport of two Na+ ions together with one amino acid molecule. The transport system has a slightly greater affinity for L-glutamate, but transports the smaller L-aspartate somewhat faster. Besides the L-isomers also D-glutamate and D-aspartate were found to depolarize the tubular cells which suggests that also the D-isomers are absorbed in the tubule, however they do not seem to use the same transport system as the L-isomers. In addition to the transport system in the brushborder, a similar Na+-dependent, rheogenic transport system for L-glutamate and L-aspartate was also found in the peritubular cell membrane, as deduced from cell cell depolarizations in response to these substrates applied peritubularly. The simultaneous presence of Na-driven transport systems in the apical and basal cell membrane which is not found with other amino acids, may explain the high intracellular accumulation of L-glutamate and L-aspartate in the kidney and provides a rational basis for explaining clinically observed cases of dicarboxylic aminoacidurias.

Insulin binding and degradation by luminal and basolateral tubular membranes from rabbit kidney.

Insulin influences certain metabolic and transport renal functions and is avidly degraded by the kidney, but the relative contribution of the luminal and basolateral tubular membranes to these events remains controversial. We studied (125)I-insulin degradation [TCA and immunoprecipitation (IP) methods] and the specific binding of the hormone by purified luminal (L) and basolateral (BL) tubular membranes. These were prepared from rabbit kidney cortical homogenates by differential and gradient centrifugation and ionic precipitation steps in sequence, which resulted in enrichment vs. homogenate of marker enzymes' activities (sodium-potassium-activated adenosine triphosphatase for BL and maltase for L) of 8- and 12-fold, respectively. Both fractions degraded insulin avidly and bound the hormone specifically without saturation even at pharmacologic concentrations (10 muM). At physiologic insulin concentrations (0.157 nM) BL membranes degraded substantial amounts of insulin (44.2+/-2.6 and 40.7+/-2.2 pg/mg protein per min by the TCA and IP methods, respectively), even though at lesser rates (P < 0.001) than the luminal fraction (67.2+/-2.3 and 75+/-6.2 pg/mg protein per min, respectively); the rate of insulin catabolism by BL membranes was significantly higher (P < 0.001) than that which could be attributed to their contamination by luminal components [12.2+/-1.9 pg/mg per min (TCA method), or 13.7+/-1.9 pg/mg per min (IP method)]. Competition experiments suggested that insulin-degrading activity in both fractions includes both specific and nonspecific components. In contrast to degradation, insulin binding by both membranes was highly specific for native insulin and was severalfold higher in BL than L membranes [17.5+/-1.3 vs. 4.5+/-0.4 fmol/mg protein (P < 0.001) at physiologic insulin concentrations]. Despite the marked difference in the binding capacity for insulin by the two membranes, the patterns of labeled insulin displacement by increasing amounts of unlabeled hormone were superimposable (50% displacement required approximately 3 nM), suggesting that their receptors' affinity for insulin was similar. These observations provide direct evidence that interaction of insulin with the kidney involves binding and degradation of the hormone at the peritubular cell membrane.

Electrophysiological analysis of rat renal sugar and amino acid transport. IV. Basic amino acids.

Electrophysiological techniques were used to study the transport of the basic amino acids L-arginine, L-lysine and L-ornithine in rat kidney proximal tubule in vivo. Tubular cells were punctured with microelectrodes and the response of the cell membrane potential to sudden applications of the amino acids was measured. In the presence of physiological Na+ concentrations luminal perfusion with millimolar concentrations of basic amino acids depolarized the tubular cells in a concentration dependent fashion by up to 15 mV, while in the absence of Na+ no significant potential changes were observed. These observations indicate that the basic amino acids are taken up into the cell across the brushborder in coupling with Na+ ions in a similar way as neutral and acidic amino acids, and that simple conductive pathways for uncoupled flow of the basic amino acids do either not exist or are quantitatively negligible in the brushborder. From kinetic measurements and competition experiments it was concluded that all basic amino acids are transported by the same transport system, which however does not accept acidic or neutral amino acids (with the possible exception of L-cystine). Perfusion of the peritubular capillaries with millimolar concentrations of basic amino acids depolarized the cells only by approximately 1 mV, both in the presence and absence of Na+. This observation may indicate that a passive uncoupled transport pathway for basic amino acids is present in the peritubular cell membrane to allow exit from cell to interstitial space, if the intracellular concentration rises high enough to overcome the cell membrane potential.

Electrophysiological analysis of rat renal sugar and amino acid transport. II. Dependence on various transport parameters and inhibitors.

Transepithelial and cellular electrical potential changes were measured in response to luminal perfusion of D-glucose and related substrates in micropuncture experiments on rat kidney in vivo. By studying the dependence of the potential response on various experimental parameters, some insight was obtained into the mechanism of Na+ coupled glucose absorption. The experiments confirm the driving forces for glucose absorption in the living cell to be: a) the Na concentration gradient, b) the electrical potential gradient and c) the glucose concentration gradient across the brush-border membrane. Furthermore they describe the substrate specificity of the cotransport mechanism and the mechanism of inhibition of D-glucose transport by various inhibitors, such as phlorizin, harmaline and ouabain. The latter experiments suggest that the active Na+ pump in the peritubular cell membrane, which establishes the Na+ ion gradient and the electrical potential gradient across the brushborder, contributes a measurable partial conductance to the overall electrical conductance of the peritubular cell membrane.

Mechanism of distal tubular chloride transport in Amphiuma kidney.

To characterize the mechanism of chloride transport across individual cell membranes, experiments were carried out on early distal tubules in the doubly perfused Amphiuma kidney and net chloride flux, transepithelial and transmembrane cell potentials, and intracellular chloride activity measured. Net chloride flux was evaluated by a modified stationary microperfusion technique, and intracellular and intraluminal chloride activities by means of double-barreled liquid ion exchange microelectrodes. Control conditions were characterized by significant net volume and chloride reabsorption, a transepithelial potential difference of +9.0 +/- 0.5 mV (lumen positive), and cell chloride activities above electrochemical equilibrium across both luminal and peritubular cell membranes. Following luminal application of furosemide (5 X 10(-5) M) or perfusion with either a sodium- or chloride-free solution, net flux of chloride fell dramatically, the transepithelial potential difference was abolished, and cell chloride activity dropped sharply to approach electrochemical equilibrium. The decrease in transepithelial potential difference was fully accounted for by hyperpolarization of the basolateral cell membrane potential. These results are consistent with a furosemide-sensitive, electrically neutral sodium chloride cotransport mechanism across the luminal cell membrane.

Peritubular interaction of unconjugated bilirubin in isolated perfused rat kidney.

1. The interaction of unconjugated bilirubin with peritubular cell membranes of the rat kidney was studied by means of an isolated rat-kidney preparation applying the multiple-indicator-dilution technique. 2. Inulin was used as an extracellular marker and p-aminohippuric acid as a model of organic anion that interacts with the peritubular membrane. 3. A single renal artery injection of a mixture containing inulin and unconjugated bilirubin was followed by the appearance of the two compounds in the venous effluent. The unconjugated bilirubin curve was always under the curve of inulin and its mean transit time was always less than that of inulin. 4. The cumulative venous recovery of inulin was higher than that of unconjugated bilirubin. 5. When unconjugated bilirubin uptake was plotted against the injected dose of pigment the relationship suggested a saturation phenomenon. 6. The recovery of p-aminohippuric acid was significantly increased when unconjugated bilirubin was added. 7. The results provide evidence for the interaction of unconjugated bilirubin with the peritubular cell membranes of rat kidney.

Oxygen requirement of bicarbonate-dependent sodium reabsorption in the dog kidney.

The ratio between changes in sodium reabsorption and renal oxygen consumption (Na/O2) was measured in anesthetized dogs at high plasma bicarbonate concentration (32 +/- 1 mM); ethacrynic acid was infused continuously to prevent variations in transcellular NaCl reabsorption when sodium reabsorption was altered by varying plasma PCO2 and glomerular filtration rate (GFR). At high plasma PCO2 (110 mmHg) sodium reabsorption varied in proportion to GRF between 50 and 125% of control GFR (glomerulotubular balance). By reducing PCO2 to 20 mmHg, sodium reabsorption was reduced by 50-60% at constant GFR. The Na/O2 ratio was not significantly different during the two procedures and averaged 48 +/- 2. The ratio between changes in NaHCO3 reabsorption and oxygen consumption averaged 17 +/- 1, which is not significantly different from the Na/O2 ratio of Na-K-ATPase-dependent sodium transport. We propose that NaHCO3 is admitted to the cell by Na+/H+ exchange and that sodium is actively transported by Na-K-ATPase across the peritubular cell membrane; NaHCO3 provides the osmotic force for paracellular reabsorption of water and NaCl (bicarbonate-dependent reabsorption) without additional energy requirement.

The effect of diffusible ions on the peritubular membrane potential of proximal tubular cells in perfused bullfrog kidneys.

Effects of extracellular diffusible ions, such as K+, Cl- and HCO3- (pH), on the peritubular membrane potential (EM) and intracellular activities of K+, (K)i, or Cl-, (Cl)i, were studied in the perfused proximal tubule of bullfrog kidneys with K+ or Cl- -selective microelectrodes. In steady-state conditions, in which both the peritubular and luminal sides were perfused with control Ringer solutions, the K+ equilibrium potential (EK) always exceeded the EM by approximately 19 mV and correlated well with the EM (correlation coefficient r = 0.78), whereas no correlation was recognized between the equilibrium potential of Cl-(ECl) and the EM. In the quick peritubular perfusion experiments, in which the extracellular diffusible ions were changed, the (K)i and (Cl)i were maintained relatively stable. The following facts were observed: (1) At constant EK, decreasing the peritubular chloride (Cl)e produced a small degree of hyperpolarization of the EM instead of depolarization. (2) At constant ECl, increasing the (K)e depolarized the EM. (3) At constant PCO2, the EM was depolarized with low HCO3- (acid) perfusions, while it was hyperpolarized with high HCO3- (alkaline) perfusions. These results are in agreement with the views that, 1) intracellular K+ in the proximal tubule is maintained by an uphill uptake mechanism on the peritubular cell membrane, (2) the ionic conductance of peritubular membrane is relatively high to K+, but low to Cl-, and (3) the pH gradient across the peritubular membrane can modulate the passive permeability to Na+ or K+.

Energetics and specificity of transcellular NaCl transport in the dog kidney

1. 1. Ouabain inhibits NaCl reabsorption and oxygen consumption in the dog kidney as expected if transport of sodium by Na-K-ATPase across the peritubular cell membrane was the only energy-requiring transport. 2. 2. The facilitated transport across the luminal cell membrane of NaCl is not specific for chloride, since ouabain and ethacrynic acid inhibit bromide and thiocyanate reabsorption as much as chloride reabsorption.

On the mechanism of HCO 3 ? permeation across the peritubular cell membrane of proximal tubular kidney cells

On the mechanism of HCO 3 ? permeation across the peritubular cell membrane of proximal tubular kidney cells

Potassium Transport in the Nephron

This review focuses on recent studies of single mammalian and amphibian tubules that aim at a delineation both of potassium transport sites and of the electrochemical driving forces across the luminal and peritubular cell membranes that control potassium translocation.

Determination of intracellular K+ activity in rat kidney proximal tubular cells.

The intracellular K+ activity of rat kidney proximal tubular cells was determined in vivo, using intracellular microelectrodes. In order to minimize damage from the impaling electrodes, separate measurements on separate cells, were performed with single-barrelled KCl-filled non-selective electrodes and single-barrelled, K+-sensitive microelectrodes, which were filled with a liquid K+-exchanger resin that has also a small sensitivity to Na+. Both electrodes had tip diameters of 0.2 μm or below. The proper intracellular localization of the electrodes was ascertained by recording the cell potential response to intermittent luminal perfusions with glucose. The membrane potential measured with the non-selective microelectrodes was −76.3±8.1 mV (n=81) and the potential difference measured with the K+-sensitive microelectrode was −7.2±5.8 mV (n=32). Based on the activity of K+ in the extracellular fluid of ∼3 mmol/l the intracellular K+ activity was estimated to be ∼82 mmol/l. Assuming equal K+-activity coefficients to prevail inside and outside the cell, this figure suggests that the intracellular K+ concentration is ∼113 mmol/l which must be considered as a lower estimate, however. The data indicate that the K+-ion distribution between cytoplasm and extracellular fluid is not in equilibrium with the membrane potential, but that K+ is actively accumulated inside the cell. This result provides direct evidence for the presence of an active K+ pump in the tubular cell membranes, which in view of other observations, must be envisaged as a (not necessarily electroneutral) Na+/K+-exchange pump which operates in the peritubular cell membrane and is eventually responsible for the major part of the tubular solute and water absorption.