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PubMed1968

Osmotic volume flow in the proximal tubule of Necturus kidney.

Volume changes due to osmotic flow in the distal portion of proximal tubules of Necturi were measured by the split oil drop technique. In agreement with previous findings no volume flow was induced by NaCl concentrations close to 60 mM. The tubule wall was found to be permeable to plasma electrolytes, which have an apparent reflection coefficient of 0.69. The mean apparent hydraulic conductivity was 0.33 x 10(-11) cm(3)/dyne sec, comparable with other epithelia. A number of lipid-insoluble nonelectrolytes of widely varying molecular size had apparent reflection coefficients of about 0.5. In view of the insensitivity to molecular size it seems likely that apparent reflection coefficients determined from tubular volume changes depend primarily on the porosity of the intercellular barrier closest to the lumen and give little information about the subsequent fate of the test substances.

Animals↗
PubMed1969

Water permeability in resting and stimulated crayfish nerve.

The hydraulic conductivity, L(p), was determined in single axons of the crayfish, Procambarus clarkii, by injecting a hypertonic sample between two drops of silicone oil and photographing the volume increase of the sample. The method has the advantage of minimizing errors due to hydrostatic pressure differences across the membrane. In resting axons an L(p) of 0.236 x 10(-8) cm/ sec per cm H(2)O was found and similar values were obtained with low external calcium concentration and when the nerve was continuously stimulated at 20-30 impulses/sec. Thus the experiments have failed to demonstrate any change of water permeability in cases in which the ionic conductance is known to change. Some possible implications of this are discussed.

Animals↗
PubMed1970

The erythrocyte ghost is a perfect osmometer.

The osmotic swelling of intact erythrocytes in hypotonic solutions was measured using microhematocrit tubes, Van Allen tubes, and a calibrated Coulter counter. In agreement with earlier workers the intact cells did not behave as perfect osmometers, the cells swelling less than predicted by the Boyle-van't Hoff law. Erythrocyte ghosts were prepared from fresh intact erythrocytes by one-step hemolysis in 0.25% NaCl at an extremely dilute concentration of cells and the membranes were sealed at 37 degrees . The ghosts were mixed with NaCl solutions of different osmolarities and the MCV (mean cell volume) of the shrunken cells immediately monitored by a calibrated Coulter counter. It was found that the MCV values of the shrunken ghosts were accurately predicted by the Boyle-van't Hoff law. These results indicate that these erythrocyte ghosts behaved as perfect osmometers.

Cell Membrane Permeability↗
PubMed1972

Temperature dependence of vasopressin action on the toad bladder.

Toad bladders were challenged with vasopressin at one temperature, fixed on the mucosa with 1% glutaraldehyde, and then subjected to an osmotic gradient at another temperature. Thus, the temperature dependence of vasopressin action on membrane permeability was distinguished from the temperature dependence of osmotic water flux. As the temperature was raised from 20 degrees to 38 degrees C, there was a substantial increase in the velocity of vasopressin action, but osmotic flux was hardly affected. In this range of temperature the apparent energy of activation for net water movement across the bladder amounted to only 1.2 kcal/mole, a value well below the activation energy for bulk water viscosity. It is suggested that osmotic water flux takes place through narrow, nonpolar channels in the membrane. When the temperature was raised from 4 degrees to 20 degrees C, both vasopressin action as well as osmotic water flux were markedly enhanced. Activation energies for net water movement were now 8.5 kcal/mole (4 degrees -9 degrees C) and 4.1 kcal/mole (9 degrees -20 degrees C), indicating that the components of the aqueous channel undergo conformational changes as the temperature is lowered from 20 degrees C. At 43 degrees C bladder reactivity to vasopressin was lost, and irreversible changes in selective permeability were observed. The apparent energy of activation for net water movement across the denatured membrane was 6.6 kcal/mole. Approximately 1 microosmol of NaCl was exchanged for 1 microl of H(2)O across the denatured membrane.

Animals↗
PubMed1985

A microscope diffusion chamber for the determination of the equilibrium and non-equilibrium osmotic response of individual cells.

A microscope diffusion chamber has been developed which allows direct observation of the dynamic osmotic response of individual cells in micro-volume suspensions. Continuous observation of stationary cells is possible including short experimental times while the extracellular chemical composition is changed. Multiple changes of solute type or concentration are easily imposed upon a single sample volume. Response times are a function of chamber configuration but response times as low as 1-10 s are possible with negligible solute concentration gradients within the sample region. The chamber is simple and economical to construct and use. It is the size of an ordinary glass microslide and it can be adapted easily to any common laboratory microscope. All standard optical techniques may be used with the chamber. Construction details and operating characteristics including important limitations are discussed. Example photomicrographs and osmotic data are included.

Animals↗
PubMed1985

Regulation of luminal membrane water permeability by water flow in toad urinary bladder.

Intramembranous particle aggregates (presumed sites for water flow) which appear in the luminal membrane consequent to ADH treatment are derived from cytoplasmic membrane structures (now termed "aggrephores") which fuse with the luminal membrane. We have previously shown that bladders stimulated in the absence of an osmotic gradient have about twice as many aggregates and about three times as many sites of aggrephore fusion as bladders stimulated with ADH in the presence of a 175 milliosmolal gradient. The present studies show that the frequency of fused aggrephores and luminal membrane aggregates can be modified as a consequence of alterations in transmembrane water flow initiated by changing the transbladder osmotic gradient during hormone stimulation. Bladders treated with ADH for 1 hr without a gradient and then for 1 hr with a gradient had approximately 1/3 as many aggregates and fusion sites as paired bladders treated for 2 hr without a gradient. Conversely, bladders treated with ADH for 1 hr with a gradient and then for 1 hr without a gradient had approximately 2x as many aggregates and fusion sites as bladders treated for 2 hr with a gradient. In other experiments we demonstrate that the time course of hormone washout is greatly accelerated if carried out in the presence of an osmotic gradient. In paired bladders that were first stimulated with ADH for 30 min in the absence of a gradient, aggregates and fusion sites as well as osmotic water permeability determined in fixed bladders, persisted at near maximum levels for 15 min of washout in the absence of a gradient.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1967

Permeability properties of Rickettsia mooseri.

The passive permeability properties of Rickettsia mooseri to both inorganic and organic solutes have been examined. Visual observations by phase-contrast microscopy of rickettsiae in macerated yolk sacs taken directly from heavily infected eggs revealed plasmolysis with hypertonic NaCl and KCl as well as with sucrose solutions. In contrast, similar visual studies of rickettsiae which had been subjected to freezing or to a purification process, or both, were plasmolyzed by hypertonic sucrose but not by hypertonic NaCl and KCl. These primary observations were extended to a variety of solutes and were placed on a quantitative basis by use of optical density and radioisotope dilution methods. Intracellular Na(+) and K(+) concentrations in processed rickettsiae, measured by flame photometry, closely paralleled the concentration of these ions in the suspending medium. It was concluded that R. mooseri appears to possess an osmotically active, functional, and structural membrane distinct from the cell wall, located at the surface of a structure analogous to the bacterial protoplast. In the intact organism, this membrane is passively impermeable to sucrose, NaCl, and KCl. However, altered permeability properties, especially to inorganic electrolytes, may be expected in rickettsiae which have been stored in the frozen state and subjected to a lengthy purification process.

Animals↗
PubMed1991

Inhibition of ion conductances by osmotic shrinkage of Madin-Darby canine kidney cells.

Osmotic swelling of Madin-Darby canine kidney (MDCK) cells enhances the ion conductances of the cell membrane, which allows release of cellular ions and subsequent regulatory cell volume decrease. The present study has been performed to test whether cell shrinkage similarly affects the ion conductances of MDCK cell membranes. Increase of extracellular osmolarity by addition of 50 mM NaCl or 100 mM mannitol leads within 3 min to a hyperpolarization of the cell membrane, a marked increase of cell membrane resistance [by 223 +/- 38% (n = 8) and 228 +/- 21% (n = 5), respectively], as well as a moderate increase of the K+ selectivity of the cell membrane (by 37 +/- 13%, n = 9). Thus exposure to hypertonic extracellular fluid decreases the cell membrane conductances including the K+ conductance. Cell volume measurements reveal a regulatory cell volume increase, which is sensitive to both furosemide and dimethylamiloride. Extracellular ATP (10 microM), which activates calcium-sensitive K+ channels, hyperpolarizes the cell membrane close to the K+ equilibrium potential. The respective values are -69.9 +/- 3.1 mV (n = 9) in isotonic fluid, -79.4 +/- 1.8 mV (n = 9) within 3 min, and -76.4 +/- 1.8 mV (n = 7) within 16-h exposure to hypertonic extracellular fluid. This observation points to a sustained increase of intracellular K+ activity after exposure to hypertonic extracellular fluid.

Adenosine Triphosphate↗
PubMed1994

Identification of a sorbitol permease in human erythrocytes.

Sorbitol, a polyol derived from glucose by the enzyme, aldose reductase, is a common organic solute in many cells. It plays a role in the osmotic regulation of epithelial cells and in the pathology of uncontrolled diabetes. To learn more about sorbitol transport, we measured D-[14C]sorbitol influx in human erythrocytes. Sorbitol influx at 37 degrees C was a linear function of sorbitol concentration over the range of 0.05-100 mM. The activation energy for sorbitol influx was 10.0 kcal/mol, and the Q10 over the range 10-50 degrees C was 1.8, higher than predicted for diffusion through an aqueous pore. Glucose transport inhibitors either had no effect (1 mM phloridzin) or minimally inhibited (approximately 35% inhibition by 10 microM cytochalasin B or 250 microM phloretin) sorbitol influx. Influx was stimulated twofold by 0.5 mM p-chloromercuribenzoic acid, an inhibitor of glucose transport, and this was reversed by 2 mM dithiothreitol. Sorbitol influx was neither Na dependent nor sensitive to changes in cell volume. Glucose, fructose, mannitol, myo-inositol, and gluconate, at four- to fivefold molar excesses over sorbitol, did not inhibit its influx. We conclude that there is a specific sorbitol transport pathway in human erythrocytes similar to the sorbitol permease in renal epithelial cells.

Animals↗
PubMed1993

Sieving of electrolytes at capillary wall of cat skeletal muscle by osmotic water flow.

To test the hypothesis that a significant proportion of transcapillary water flow occurs through solute-restricting channels, we investigated the effects of transcapillary water movement on plasma electrolytes in isolated perfused cat skeletal muscle. The lower hindlimbs of anesthetized cats were perfused with a plasma-albumin solution and were weighed to determine transcapillary water movement. Osmolality was increased 60-70 mosmol/kgH2O with sucrose, creating water fluxes of 8-10 ml.min-1.100 g-1, and the changes in the venous concentrations of sodium, potassium, and chloride were determined. The ion concentrations were all reduced by 6-7% with no significant difference between them. The amount of reduction was quantitatively explained by the flow of ion-free water from the interstitial space into plasma and the diffusion of electrolyte in the same direction. These findings support the hypothesis that important water-only transcapillary channels exist in mammalian skeletal muscle. The observations may also explain some of the electrolyte changes seen in intense exercise.

Animals↗
PubMed1985

Evidence for transcellular osmotic water flow in rat proximal tubules.

To determine the predominant pathway for transepithelial osmotic water flow, the transepithelial osmotic water permeability [Pf(TE)] and the apparent dimensions of paracellular pores and slits were determined in rat proximal convoluted tubules microperfused in vivo. To measure Pf(TE), tubules were perfused with a hyposmotic, cyanide-containing solution. Pf(TE), calculated from the observed volume flux in response to the measured log mean osmotic gradient, was 0.12-0.15 cm/s, assuming sigmaNaCl equal to 1.0-0.7, respectively. The dimensions of the paracellular pathways were determined using measured sucrose and mannitol permeabilities (nonelectrolytes confined to the extracellular space). These were 0.43 and 0.87 X 10(-5) cm/s, respectively. By using the ratio of these permeabilities, their respective free solution diffusion coefficients and molecular radii, and the Renkin equation, the radius of the nonelectrolyte-permeable pores and the total pore area/cm2 surface area/channel length were calculated to be 1.4 nm and 3.56 cm-1, respectively. Similar calculations for slits yielded a slit half-width of 0.8 nm and a total slit area/cm2 surface area/channel length of 3.16 cm-1. The osmotic water permeability of these nonelectrolyte-permeable pathways was calculated by Poiseuille's law to be 0.0018 cm/s (pores) or 0.0014 cm/s (slits), at most 2% of Pf(TE). We conclude that the nonelectrolyte-permeable pathway in the tight junctions is not the major route of transepithelial osmotic water flow in the rat proximal tubule.

Animals↗
PubMed1974

Effects of ethacrynic acid on the isolated collecting tubule.

Effects of the diuretic ethacrynic acid on osmotic water permeability were investigated in the isolated perfused collecting tubule of the rabbit kidney. The base-line water permeability of the collecting tubule was not affected when the drug (10(-4)M) alone was added to the bathing medium. Vasopressin alone in the bathing medium (2, 5 muU/ml) elicited a significant increase in osmotic water absorption. With vasopressin kept in the bathing medium, the addition of 10(-5)M ethacrynic acid depressed the hydro-osmotic effect of vasopressin by 50%. This inhibitory effect of low concentrations of ethacrynic acid could be surmounted by high, supramaximal dosage levels of vasopressin. When 10(-4)M ethacrynic acid was added to the bathing medium before vasopressin, the hydro-osmotic effect of vasopressin and the diuretic in combination was insignificant.Dibutyryl adenosine 3'5'-monophosphate (10(-4)-10(-2)M) alone in the bathing medium significantly increased baseline osmotic water flow, mimicing the effect of antidiuretic hormone. When ethacrynic acid was added together with the nucleotide, the permeability remained at the same high level. Theophylline, like the nucleotide and vasopressin, produced a significant hydro-osmotic effect. The magnitude of this response was not affected by further addition of ethacrynic acid (10(-4)M). It was concluded that ethacrynic acid is an antagonist of antidiuretic hormone. The antagonism probably occurs at the level of the receptor site of the hormone on the peritubular membrane. Antagonism to circulating antidiuretic hormone may therefore be one of the factors involved in the loss of renal concentrating ability brought about by ethacrynic acid diuresis.

Animals↗
PubMed1984

Functional characterization of the alpha adrenergic receptor modulating the hydroosmotic effect of vasopressin on the rabbit cortical collecting tubule.

To characterize the type of alpha adrenergic receptor, the effects of specific alpha adrenergic agonists and antagonists on antidiuretic hormone [( Arg8]-vasopressin [AVP])-induced water absorption were evaluated in cortical collecting tubules isolated from the rabbit kidney and perfused in vitro. In the presence of AVP (100 microU/ml), net fluid volume absorption (Jv, nanoliters per minute per millimeter) was 1.39 +/- 0.09 and osmotic water permeability coefficient (Pf, X 10(-4) centimeters per second) was 150.2 +/- 15.0. The addition of 10(-6) M phenylephrine (PE), an alpha adrenergic agonist, resulted in a significant decrease in Jv and Pf to 0.72 +/- 0.11 (P less than 0.005) and 69.9 +/- 10.9 (P less than 0.005). The addition of 10(-4) M prazosin (PZ), an alpha adrenergic antagonist, did not cause any significant change in Jv and Pf, which were 0.71 +/- 0.09 (P = NS vs. AVP + PE) and 67.8 +/- 9.5 (P = NS vs. AVP + PE), respectively. In a separate group of tubules, in the presence of AVP (100 microU/ml) and PE (10(-6) M), Jv and Pf were 0.78 +/- 0.17 and 76.1 +/- 18.0, respectively. The addition of 10(-6) M yohimbine (Y), an alpha 2 adrenergic antagonist, resulted in a significant increase in Jv to 1.46 +/- 0.14 (P less than 0.01) and Pf to 157.5 +/- 22.3 (P less than 0.005). Y (10(-4) M) or PZ (10(-4) M) alone did not significantly affect Jv and Pf in the presence of AVP )100 microU/ml). The effect of the natural endogenous catecholamine norepinephrine (NE) on Jv and Pf in the presence of AVP and propranolol (PR) was next examined. Jv and Pf were 1.53 +/- 0.07 and 176.3 +/- 5.2, respectively, in the presence of AVP (100 microU/ml) and PR (10(-4) M). The addition of NE (10(-8) M) resulted in a significant decrease in Jv to 1.19 +/- 0.11 (P less than 0.05) and Pf to 127.0 +/- 11.3 (P less than 0.02). Increasing the concentration of NE to 10(-6) M resulted in a further decrease in Jv and Pf to 0.70 +/- 0.10 (P less than 0.01 vs. NE 10(-8) M) and 68.5 +/- 10.6 (P less than 0.01 vs. NE 10(-8) M), respectively. The inhibitory effect of NE on AVP-induced water absorption was blocked by Y, but not by PZ. The effect of the alpha 2 adrenergic agonist clonidine (CD) on Jv and Pf was also examined. In the presence of AVP (10 microU/ml) Jv and Pf were 1.65 +/- 0.04 and 175.1 +/- 13.1, respectively. The addition of CD (10(-6) M) resulted in a significant decrease in Jv to 1.08 +/- 0.12 (P < 0.01) and Pf to 108.1 +/- 15.4 (P < 0.01). Increasing the concentration of CD to 10(-4) M resulted in a further significant decrease in Jv and Pf to 0.57 +/- 0.13 (P < 0.02 vs. CD 10(-6) M) and 54.7 +/- 13.8 (P < 0.01 vs. CD 10(-6) M), respectively. Similar results were obtained in the presence of AVP (100 microU/ml). The inhibitory effect of CD on AVP-induced water absorption was blocked by Y. CD did not significantly affect Jv and Pf in the presence of 8-bromo adenosine 3',5'-cyclic monophosphate. These studies indicate that alpha adrenergic agonists directly inhibit AVP-mediated water absorption at the level of renal tubule, an effect that can be blocked by specific alpha2 adrenergic antagonists, but not by specific alpha1 adrenergic antagonists. Alpha2 adrenergic stimulation directly inhibits AVP-mediate water absorption at the level of the tubule, an effect that can be blocked by a specific alpha2 adrenergic antagonist. This effect appears to be exerted at the level of activation of adenylate cyclase, since it is absent in the present of cyclic AMP.

Absorption↗
PubMed1998

Importance of interfacial water in permeabilization of ceramide bilayers.

The role of structured water at the membrane-aqueous interface in regulating the water permeability functions of the epidermal horny layer (Stratum corneum) is not clear at present. The subject of this review is the effect of perturbed interfacial water structure on the relative water permeability properties in shrinkage experiments of reconstituted vesicle suspensions of ceramides and cholesterol in the gel phase. Water structure was perturbed by a series of reagents known to affect the phase equilibrium of lipid assemblies. Multilamellar bilayers containing synthetic ceramides N-palmitoyl-D-L-dihydrosphingosine, and N-oleoyl-D-sphingosine were used as model membranes. The reagent binding to the ceramide membrane was determined by fluorescence measurements with 1-anilino, 8-naphtalene sulphonate (ANS). A dependence of number of binding sites for ANS on the type of reagent was found. The change of relative water permeability with concentration of reagents was studied. Analysis of these data reveals that the anomalous behaviour of relative water permeability with the concentration of reagents used is due to concentration-dependent structural transitions of the bound water at the membrane-aqueous interface. Different possibilities of interaction of the reagents with the interface are discussed. At the end, it is suggested that the interfacial water in multilamellar ceramide structures may also contribute to the water holding and permeability barrier functions of cell membrane complex within the stratum corneum.

Animals↗