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G Giebisch

Publications and source records attributed to G Giebisch.

At least 127 records · Page 7Linked to original sources
PubMed1990

Distal perfusion studies: transport stimulation by native tubule fluid.

It is well established that potassium secretion into the distal tubule increases with the rate of flow. In a previous study [G. Malnic, R. W. Berliner, and G. Giebisch. Am. J. Physiol. 256 (Renal Fluid Electrolyte Physiol. 25): F932-F1271, 1989] we found that the increase with the rate of perfusion with a fluid made up to resemble that normally found in the early distal tubule was substantially less than the increase in free-flow conditions [R. N. Khuri, M. Wiederholt, N. Strieder, and G. Giebisch. Am. J. Physiol. 228: 1249-1261, 1975]. Because of the possibility that some important component was missing from the artificial fluid, we have carried out another series of experiments in which distal tubules were perfused with fluid collected from late proximal tubules and compared the results with those obtained when tubules were perfused with an artificial fluid with an electrolyte composition similar to that of late proximal fluid. When proximal tubule fluid was used, the potassium concentrations in the collected distal fluid were higher and better maintained with increasing flow than when the artificial fluid was used, and consequently the rate of potassium secretion was substantially greater with the proximal tubule fluid, approaching the results of previous studies in free flow. The nature of the component missing from the artificial solution is not known.

Absorption↗
PubMed1990

Morphological and physiological responses to aldosterone: time course and sodium dependence.

The time course and Na+ dependence of morphological responses to a physiological elevation of aldosterone were examined in the renal cortical collecting duct. Male Sprague-Dawley rats were adrenalectomized and given a basal replacement infusion of aldosterone and dexamethasone by osmotic minipump. The animals were given additional aldosterone by either intravenous infusion or by a second implanted osmotic minipump for either 5 h or 1, 3, 7, or 14 days. Animals were fed either a Na(+)-replete or a low-Na+ diet. After these treatments we conducted a morphometric analysis on kidneys processed for electron microscopy by standard techniques. Increased aldosterone induced detectable proliferation of the basolateral membrane of principal cells after 24 h. Basolateral membrane continued to increase rapidly through 3 days of exposure and thereafter at a modest rate. Cell area and the surface density of the basolateral membrane were also increased by aldosterone treatment. The morphological response to aldosterone was markedly inhibited by maintaining animals on a low-Na+ diet. Acute exposure to elevated aldosterone levels (5 h) increased K+ secretion by principal cells without alterations in basolateral membrane area. Chronic (greater than 24 h) exposure to aldosterone induces morphological adaptations that are accompanied by further elevations in K+ secretion.

Aldosterone↗
PubMed1990

Regulation of small-conductance K+ channel in apical membrane of rat cortical collecting tubule.

We used the patch-clamp technique to study the activity and regulation of single potassium channels in the apical membrane of isolated cortical collecting tubules (CCT) of rat kidney. With 140 mM K+ in the pipette the inward conductance of the channel in cell-attached patches at 37 degrees C was 35 pS (n = 106, NaCl-Ringer or 70 mM KCl and 70 mM NaCl in the bath), and the outward conductance was 15 pS (n = 15, 70 mM NaCl + 70 mM KCl in the bath). Mean open probability (Po) of the channel is voltage independent and 0.96 (n = 106). The channel displayed one open state with a mean lifetime of 18.6 ms and one closed state with a mean lifetime of 0.7 ms (n = 20). Selectivity ratio between K+ and Na+ is 20 (n = 5). High-potassium diet increased channel incidence from control 32% (53 patches with channel from 165 patches) to 64% (53 patches with channels from 83 patches). The channel could be blocked by 1 mM Ba2+ (n = 7, Ba2+ in the pipette); however, 5 mM tetraethylammonium (n = 9, TEA in the pipette) did not block the channel activity. The channel was very sensitive to intracellular pH (n = 6). Changing bath pH facing cytoplasmic side of inside-out patches from 7.4 to 6.9 reversibly reduced Po from 0.9 to 0.1. Addition of 1 mM ATP (n = 7) to bath almost completely inhibited channel activity in inside-out patches. This ATP-induced inhibition was fully reversible and was found to be dependent on the ratio of ATP to ADP, since adding 0.5 mM ADP to bath solution relieved the ATP-induced blockade. Results indicate that intracellular pH, concentration of ATP, and ratio of ATP to ADP are important regulators of potassium channel activity in the apical membrane of rat CCT, and the properties of the channel make it a strong candidate for K+ secretion in this nephron segment.

Adenosine Triphosphate↗
PubMed1989

Mechanism of aldosterone-induced increase of K+ conductance in early distal renal tubule cells of the frog.

Isolated early distal tubule cells (EDC) of frog kidney were incubated for 20-28 hr in the presence of aldosterone and then whole-cell K+ currents were measured at constant intracellular pH by the whole-cell voltage-clamp technique. Aldosterone increased barium-inhibitable whole-cell K+ conductance (gK+) threefold. This effect was reduced by amiloride and totally abolished by ouabain. However, aldosterone could still raise gK- in ouabain-treated cells in the presence of furosemide. We tested whether changes in intracellular pH (pHi) could be a signal for cells to regulate gK+. After removal of aldosterone, the increase in gK+ was preserved by subsequent incubation for 8 hr at pH 7.6 but abolished at pH 6.6. In the complete absence of aldosterone, incubation of cells at pH 8.0 for 20-28 hr raised pHi and doubled gK+. Using the patch-clamp technique, three types of K+-selective channels were identified, which had conductances of 24, 45 and 59 pS. Aldosterone had no effect on the conductance or open probability (Po) of any of the three types of channels. However, the incidence of observing type II channels was increased from 4 to 22%. Type II channels were also found to be pH sensitive, Po was increased by raising pH. These results indicate that prolonged aldosterone treatment raises pHi and increases gK+ by promoting insertion of K+ channels into the cell membrane. Channel insertion is itself triggered by raising both pHi and increasing the activity of the Na+/K+ pump in early distal cells of frog kidney.

Aldosterone↗
PubMed1989

Studies on the mechanism of rubidium-induced kaliuresis.

Renal clearance and electron microprobe methods were used 1) to elucidate the effects of chronic rubidium administration on potassium transport and 2) to localize, by the use of amiloride in acute experiments, the tubule site of interaction between rubidium and potassium. Substitution of drinking water by a 50 mM rubidium chloride solution for 9 to 11 days led to significant hypokalemia (plasma potassium 2.5 +/- 0.1 mM; plasma potassium plus rubidium 3.3 +/- 0.1 mM). Compared to a control group (reduction of plasma potassium to 3.4 +/- 0.1 mM by short-term potassium depletion) with a fractional potassium excretion of 2.1 +/- 0.3%, rubidium-treated rats excreted potassium at a much higher rate of 14.6 +/- 3.0%. The potassium content of principal cells was, however, significantly lower in rubidium-treated than in potassium-deprived animals. Similar to experiments in which rubidium was given acutely (3 hours), chronic rubidium administration was associated with preferential accumulation of rubidium in all tubule cells relative to potassium. Rubidium clearances were uniformly below those of potassium. Amiloride abolished the difference between rubidium and potassium clearances and sharply reduced the excretion of both cations. In view of the known site of action of amiloride, this suggests a distal tubule site of rubidium action on potassium transport. Amiloride also reduced or abolished the preferential uptake of rubidium into all but intercalated tubule cells. Marked cell heterogeneity of rubidium accumulation into intercalated cells was observed: One subpopulation, with low cell chloride, retained rubidium more effectively than another subpopulation with high cell chloride.

Amiloride↗
PubMed1989

Na-K pump current in the Amphiuma collecting tubule.

There is strong evidence supporting the hypothesis of an electrogenic Na-K pump in the basolateral membrane of several epithelia. Thermodynamic considerations and results in nonepithelial cells indicate that the current carried by the pump could be voltage dependent. In order to measure the pump current and to determine its voltage dependence in a tight epithelium, we have used the isolated perfused collecting tubule of Amphiuma and developed a technique for clamping the basolateral membrane potential (Vbl) through transepithelial current injection. The transcellular current was calculated by subtracting the paracellular current (calculated from the transepithelial conductance measured in the presence of luminal amiloride) from the total transepithelial current. Basolateral membrane current-voltage (I-V) curves were obtained in conditions where the ratio of the pump current to the total basolateral membrane current had been maximized by loading the cells with Na+ (exposure to low-K+ bath), and by blocking the basolateral K+ conductance with barium. The pump current was defined as the difference of the current across the basolateral membrane measured before and 10-15 s after the addition of strophanthidin (20 microM) to the bath solution. With a bath solution containing 3 mM K+, the pump current was nearly constant in the Vbl range of -20 to -80 mV (52 +/- 5 microA.cm-2 at -60 mV) but showed a marked voltage dependence at higher negative Vbl (pump current decreased to 5 +/- 9 microA.cm-2 at -180 mV). In a 1.0 mM K bath, the shape of the pump I-V curve was similar but the amplitude of the current was decreased (24 +/- 4 microA.cm-2 at -60 mV). In a 0.1 mM K bath, the pump current was not significantly different from 0. Our results indicate that the basolateral Na-K pump generates a current which depends on the extracellular potassium concentration. With physiological peritubular concentration of K+ and in the physiological range of potential, the pump activity, measured as the pump-generated current, was independent of the membrane potential.

Animals↗
PubMed1989

Ultrastructure of Amphiuma distal nephron: evidence for cellular heterogeneity.

To obtain more information on the ultrastructure of the distal nephron of the salamander, Amphiuma, we conducted freeze-fracture electron microscopy and morphometric experiments. In the early distal tubule, the organization of the tight junction is variable, containing from one to two strands in the proximal region and four strands in distal regions. The length density of the tight junction in this segment varies from greater than 60 m/cm2 of apical membrane surface to less than 10/cm2 of apical membrane surface. These observations agree with a previous study demonstrating that the junction of this segment exhibits considerable axial heterogeneity. The junctions of the late distal tubule and collecting tubule are more complex. In the late distal tubule, the tight junction is composed of 6-8 strands, whereas the tight junction of the collecting tubule is composed of 8-12 strands. The collecting tubule contains principal cells and two types of intercalated cells: alpha and beta. The alpha-cells contain a high density of rod-shaped particles in the apical plasma membrane and in membranes of apical cytoplasmic vesicles. The beta-cells contain rod-shaped particles only in the basolateral membrane. In principal cells, we observed a novel organization of intramembranous particles within the apical plasma membrane. A model describing the relationship of the two types of intramembranous particles within the membrane is presented. This study demonstrates that the amphibian and mammalian distal nephron share many morphological characteristics including cellular and axial heterogeneity.

Animals↗
PubMed1989

K activity of CCD principal cells from normal and DOCA-treated rabbits.

We used liquid ion exchanger and conventional microelectrodes to evaluate the effects of mineralocorticoids on the intracellular K activity (aiK) and K transport properties of principal cells (PC) of isolated cortical collecting ducts (CCDs). Hoffman modulation optics and electrophysiological methods were used to identify PC. K activity was measured with two single-barreled electrodes. We found that aiK of PC from deoxycorticosterone acetate (DOCA)-treated rabbits (97.6 mM) was not different from controls (94.8 mM). The driving forces for K transport across the basolateral membrane favored cell to bath (reabsorption) in PCs from controls and bath to cell (secretion) in PCs from DOCA-treated rabbits. However, the driving force for K secretion across the apical membrane was not significantly different between the two groups. We used the intracellular aiKs and bath ion substitutions (gluconate for Cl and K for Na) to evaluate the effects of DOCA on the ion-selective properties of the basolateral membrane of PC. DOCA increased PK/PCl from 0.33 to 0.89. Our conclusion was as follows: in PC of control rabbits K is above electrochemical equilibrium across the basolateral membrane. However, the basolateral K conductance is probably too small for significant K recycling. In PC of DOCA-treated rabbits the aiK is below electrochemical equilibrium across the basolateral membrane and the K conductance is increased. These effects enhance K secretion across this border while maintaining cell K constant.

Algorithms↗
PubMed1989

Flow dependence of K+ secretion in cortical distal tubules of the rat.

Superficial distal tubules were pump perfused (range 0-35 nl/min), generally with solutions similar in composition to early distal tubule fluid, in control, K+-depleted, acutely K+-loaded, and K+-adapted rats with the use of double-barreled resin-reference microelectrodes to measure K+ and Na+ activities and transepithelial potential differences (PD). When perfusion rate decreased from 35 to 5 nl/min in control animals, [K+] increased from 2 mM to between 10 and 20 mM, remaining at these levels as perfusion rate was decreased further. In low-K+ rats, the change in K+ activity with flow was greatly attenuated. In K+-loaded and K+-adapted rats, [K+] was higher than in controls at all flow rates. Na+ concentrations and lumen-negative PD increased with high flow rates in control rats. Addition of 10(-3) M amiloride blocked the increase in luminal K+ with low flow rates. In the physiological range of late distal flow rates, luminal [K+] remains constant and similar to the concentration attained in the steady state. At higher flow rates, [K+] declines, and K+ balance, PD, and Na+ reabsorption modulate the relationship between K+ secretion and flow rate.

Amiloride↗
PubMed1989

Effects of acetate on luminal acidification processes in the S3 segment of the rabbit proximal tubule.

We determined that, in the nominal absence of HCO-3, acetate (Ac-) doubles luminal acidification in the S3 segment of the rabbit proximal tubule. This stimulation had two components, one that was dependent on Na+ and luminal Ac- and a second that was independent of Na+ but dependent on basolateral Ac-. In the presence of 25 mM HCO-3, Ac- did not stimulate acid secretion (i.e., HCO-3 reabsorption), but actually inhibited it. The inhibition was 35% with bilateral Ac- and 15% with basolateral Ac-. The effects of Ac- were reversible both in the absence and presence of HCO-3, and are present at concentrations as low as 1 mM. We conclude that acetate (i.e., monocarboxylates) has a significant effect on luminal acidification processes both in the presence and absence of HCO-3.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1989

Reflection coefficients and water permeability in rat proximal tubule.

Simultaneous microperfusion of proximal tubules and peritubular capillaries in kidneys of rats anesthetized with Inactin was used to measure reflection coefficients. All perfusates contained cyanide to inhibit active transport; the tubular perfusate was isotonic and the peritubular capillaries were perfused with solutions made hypertonic with NaCl, NaHCO3, L-glucose, or sodium ferrocyanide. Measurements of recollected fluid enabled a precise mean gradient and ionic fluxes to be calculated; net water flux was measured with inulin. Imposed gradients always partly dissipated. Reflection coefficients were 0.59 +/- 0.01 for NaCl, 0.87 +/- 0.04 for NaHCO3-, and 0.96 +/- 0.07 for ferrocyanide, assuming that L-glucose was 1. Water permeability of the proximal tubule was 1,030 microns/s. Ionic permeability of Cl- (21.6 +/- 1.3 X 10(-5) cm/s) was greater than that for Na+ (13.3 +/- 2.7 X 10(-5) cm/s); permeability for L-glucose was 5.4 +/- 1.3 X 10(-5), and for ferrocyanide ions 2.7 +/- 0.9 X 10(-5) cm/s. It is concluded that in rat proximal tubule both NaCl and NaHCO3 have reflection coefficients less than 1.0 and solute asymmetry across the epithelium is a significant driving force for fluid reabsorption. Furthermore the data suggest that there is a significant contribution of solvent drag to solute movement.

Animals↗
PubMed1989

Osmotic forces driving water reabsorption in the proximal tubule of the rat kidney.

Simultaneous microperfusion of proximal tubules and peritubular capillaries in kidneys of rats anesthetized with Inactin was used to measure reabsorption of fluid in response to an imposed osmotic gradient. The tubular fluid was isotonic and the peritubular capillaries were made hypertonic with NaCl or NaHCO3. Mean gradients and ionic fluxes were measured. When no gradient was imposed tubular fluid became hypotonic and rate of fluid reabsorption was 0.700 nl.mm-1.min-1. Imposition of a 25 mM NaCl gradient increased fluid flux to 3.887 nl.mm-1.min-1, whereas 25 mM NaHCO3 stimulated 5.226 ml/mm fluid reabsorption. This gave a relative reflection coefficient of NaCl:NaHCO3 of 0.73. Apparent water permeability varied with highest values for the smallest gradients. This suggests the possibility of a compartment in the epithelium that is hypertonic to the peritubular capillaries. The hypertonicity required to account for fluid movement was 6-16 mosmol/kg.

Absorption↗
PubMed1989

Basolateral sodium-coupled acid-base transport mechanisms of the rabbit proximal tubule.

We studied Na+-coupled acid-base transport at the basolateral membrane of single, isolated, perfused rabbit proximal tubules by monitoring the time course of intracellular pH (pHi). The latter was determined using a microspectrofluorometric apparatus to alternatively excite the pH-sensitive fluorescent dye 2',7'-bis-2-carboxyethyl-5(and -6)-carboxyfluorescein (BCECF) at 440 and 490 nm, while the fluorescence emission, was measured at 530 nm. All experiments were conducted in the nominal absence of HCO-3 S1, S2, and S3 segments from both superficial and juxtamedullary nephrons were examined individually. We found that removing Na+ from both the lumen and bath (i.e., basolateral solution) caused pHi to fall from 7.24 to 6.75 in the superficial S1 segment (SS1), from 7.14 to 6.67 in the SS2, and from 7.09 to 6.69 in the SS3. Similarly, in juxtamedullary nephrons (J), bilateral Na+ removal caused pHi to fall from 7.25 to 6.76 in the JS1, from 7.16 to 6.71 in the JS2, and from 7.10 to 6.75 in the JS3. In all six proximal tubule subtypes, returning Na+ to the bath caused pHi to recover (i.e., increase). 4,4'-Diisothiocyanostilbene-2,2'-disulfonic acid (DIDS, 50 microM), an inhibitor of HCO-3 transport systems, blocked this Na+-dependent pHi recovery in all three superficial subtypes and the JS3 but had no effect in either the JS1 or JS2. On the other hand, 50 microM ethylisopropyl amiloride (EIPA), an inhibitor of Na-H exchange, blocked the Na+-dependent pHi recovery in the JS1 and JS2 but had no effect in the JS3 or any of the superficial subtypes.(ABSTRACT TRUNCATED AT 250 WORDS)

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1989

Renal bicarbonate reabsorption in the rat. III. Distal tubule perfusion study of load dependence and bicarbonate permeability.

Using continuous microperfusion techniques, we studied the load dependence of bicarbonate reabsorption along cortical distal tubules of the rat kidney and their bicarbonate permeability. Net bicarbonate transport was evaluated from changes in tracer inulin concentrations and total CO2 measurements by microcalorimetry. Bicarbonate permeability was estimated from the flux of total CO2 along known electrochemical gradients into bicarbonate-and chloride-free perfusion solution containing 10(-4) M acetazolamide. Transepithelial potential differences were measured with conventional glass microelectrodes. Significant net bicarbonate reabsorption occurred at luminal bicarbonate levels from 5 to 25 mM, and at perfusion rates from 5 to 30 nl/min. Bicarbonate reabsorption increased in a load-dependent manner, both during increments in luminal bicarbonate concentration or perfusion rate, reaching saturation at a load of 250 pmol/min with a maximal reabsorption rate of approximately 75 pmol/min.mm. Rate of bicarbonate reabsorption was flow dependent at luminal concentrations of 10 but not at 25 mM. During chronic metabolic alkalosis, maximal rates of reabsorption were significantly reduced to 33 pmol/min.mm. The bicarbonate permeability was 2.32 +/- 0.13 x 10(-5) cm/s in control rats, and 2.65 +/- 0.26 x 10(-5) cm/s in volume-expanded rats. Our data indicate that at physiological bicarbonate concentrations in the distal tubule passive bicarbonate fluxes account for only 16-21% of net fluxes. At high luminal bicarbonate concentrations, passive bicarbonate reabsorption contributes moderately to net reabsorption of this anion.

Absorption↗
PubMed1988

Na+ and K+ fluxes stimulated by Na+-coupled glucose transport: evidence for a Ba2+-insensitive K+ efflux pathway in rabbit proximal tubules.

Addition of glucose or the nonmetabolizable analogue alpha-methyl-D-glucoside to rabbit proximal tubules suspended in a glucose- and alanine-free buffer caused a sustained increase in intracellular Na+ content (+43 +/- 7 nmol.(mg protein)-1) and a concomitant but larger decrease in K+ content (-72 +/- 11 nmol.(mg protein)-1). A component of the net K+ efflux was Ba2+ insensitive, and was inhibited by high (1 mM) but not low (10 microM) concentrations of the diuretics furosemide and bumetanide. The increase in intracellular Na+ content is consistent with the view that the increased rates of Na+ and water transport seen in the proximal tubule in the presence of glucose can be attributed (at least in part) to a stimulation of basolateral pump activity by an increased [Na+]i.

3-O-Methylglucose↗
PubMed1988

Voltage dependence of the basolateral membrane conductance in the Amphiuma collecting tubule.

The basolateral potassium conductance of cells of most epithelial cells plays an important role in the transcellular sodium transport inasmuch as the large negative equilibrium potential of potassium across this membrane contributes to the electrical driving force for Na+ across the apical membrane. In the present study, we have attempted to establish the I-V curve of the basolateral membrane of the Amphiuma collecting tubule, a membrane shown to be K+ selective. Transepithelial I-V curves were obtained in short, isolated perfused collecting tubule segments. The "shunt" conductance was determined using amiloride to block the apical membrane Na+ conductance. In symmetrical solutions, the "shunt" I-V curve was linear (conductance: 2.2 +/- 0.3 mS.cm-2). Transcellular current was calculated by subtracting the "shunt" current from the transepithelial current in the absence of amiloride. Using intracellular microelectrodes, it was then possible to measure the basolateral membrane potential simultaneously with the transcellular current. The basolateral conductance was found to be voltage dependent, being activated by hyperpolarization: conductance values at -30 and -80 mV were 3.6 +/- 1.0 and 6.6 +/- 1.0 mS.cm-2, respectively. Basolateral I-V curves were thus clearly different from that predicted by the "constant field" model. These results indicate that the K+-selective basolateral conductance of an amphibian collecting tubule shows inward ("anomalous") rectification. Considering the electrogenic nature basolateral Na-K-pump, this may account for coupling between pump-generated potential and basolateral K+ conductance.

Animals↗
PubMed1988

Renal excretion of rubidium and potassium: an electron microprobe and clearance study.

A combination of clearance and electron microprobe studies was carried out to investigate renal rubidium excretion and rubidium distribution between plasma and individual tubule cells. Saline-infused animals were compared with potassium-loaded rats and another group in which rubidium was given in such amounts that the sum of plasma rubidium plus potassium equalled the potassium concentration in the potassium-loaded rats. The renal clearance of rubidium was uniformly less than that of potassium. Nevertheless, rubidium stimulated fractional potassium excretion above the levels observed in both saline- and potassium-loaded animals. When compared with their plasma concentrations, rubidium was concentrated in all tubule cell types more than potassium, and this is most likely due to restriction of passive diffusion of rubidium from cells to extracellular fluid. In addition, heterogeneity of intercalated cell ion composition was observed: one cell group had high chloride and potassium, but low rubidium contents, whereas the other was characterized by low chloride and potassium, but high rubidium contents.

Animals↗