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Osmotic water permeability of rabbit corneal endothelium and its dependence on ambient concentration.

We measured the fluid flow osmotically induced by sucrose concentration differences across the isolated rabbit corneal endothelium to determine if its osmotic water permeability, Pos, depends on the concentrations on both sides as well as on the concentration difference across that tissue. We found that when the osmotically induced fluid flow went from stroma to aqueous, Pos decreased from 35 +/- 4 mum/s to 20 +/- 3 mum/s when an additional 20 mosM sucrose was added to the solutions on both sides of the endothelium. However, when the osmotically induced fluid flow was towards the stroma, Pos remained practically unchanged. (28 +/- 4 mum/s vs. 31 +/- 5 mum/s), when additional 20 mosM sucrose was present on both sides. These changes in the measured permeability are consistent with the possibility that sucrose would be swept into the intercellular channels by the osmotically induced fluid flow. We also confirmed that an osmotic gradient can 'prime' the fluid pump. After a gradient was removed, gradients which had previously induced flow from stroma to aqueous caused an increase in the basal fluid pump rate, while gradients in the opposite direction decreased that fluid pump rate.

Animals↗

Osmotic stabilizers differentially inhibit permeability alterations induced in Vero cells by Clostridium perfringens enterotoxin.

Using a sensitive Vero (African green monkey kidney) cell model system, studies were performed to further investigate whether Clostridium perfringens enterotoxin acts via disruption of the colloid-osmotic equilibrium of sensitive cells. Enterotoxin was shown to cause a rapid loss of intracellular 86Rb+ (Mr approx. 100) with time- and dose-dependent kinetics. The enterotoxin-induced release of intracellular 86Rb+ preceded the loss of two larger labels, 51Cr label (Mr approx. 3500) and 3H-labeled nucleotides (Mr less than 1000). The osmotic stabilizers, sucrose and poly(ethylene glycol), differentially inhibited enterotoxin-induced larger label loss versus 86Rb+ loss. Further, enterotoxin was shown to cause a rapid influx of 24Na+ that was not significantly inhibited by osmotic stabilizers. Additional studies demonstrated that lysosomotropic agents were not protective against characteristic enterotoxin-induced membrane permeability alterations or morphological damage. Taken collectively, these results are consistent with an action for enterotoxin which involves a disruption of the osmotic equilibrium.

Animals↗

Permeability of cultured megakaryocytopoietic cells of the rat to dimethyl sulfoxide.

The permeability of the membrane of the rat megakaryocytopoietic cell to dimethyl sulfoxide was measured to assess its availability to the intracellular compartment. The method used was osmotic, and measured the initial loss of cell water followed by a reswelling to isotonic volume when cells were placed in culture media containing 0.6 M DMSO. Values for the hydraulic coefficient, Lp, the permeability of the membrane to DMSO, wRT , and the reflection coefficient were calculated from the equations of Kedem and Katchalsky . The average value at 25 degrees C for Lp was 0.46 micron min-1 atm1 ; wRT was 9.3 micron min-1, and the reflection coefficient was 0.65. At these cell volumes, 50% equilibration occurred in 5 sec. Cells equilibrated in 0.6 M DMSO increased their volume of osmotically inactive water. Coupled with this phenomenon of stabilization of water was a reduction in the hydraulic coefficient by 50%. These findings are discussed in the context of current hypotheses about cellular viability during freezing and thawing in the presence and absence of cryoprotectants.

Animals↗

Determination of fluid extraction and osmotic conductance sigma K in the lung with hypertonic NaCl infusion. II. Experiments.

The change in venous blood density in an in vitro rabbit lung preparation was measured after the osmolarity of the blood was elevated 17 to 50 mosmol/liter by a constant arterial infusion of hypertonic saline. We observed a transient density decrease of the blood flowing from the lung and then a return to the preinfusion density within 10 sec, an indication of the rapid completion of fluid extraction from the interstitia by the elevation in osmotic pressure. By fitting the time course of the density change with the prediction of an extraction model, we obtained the osmotic conductance sigma K (the product of the reflection and filtration coefficient) of the lung due to the increase in NaCl osmotic pressure as 1.33 +/- 0.18 ml/[hr.(mosmol/liter).g wet lung tissue] (mean +/- SEM), the interstitial volume participating in the extraction process as 0.27 +/- 0.04 ml/g of lung tissue, and the volume of fluid extracted as 1.12 +/- 0.16 microliter/g tissue for every mosmol/liter elevation. Since we also found no significant difference between the osmotic extraction parameters determined in blood-perfused lungs and those determined in plasma-perfused lungs, we concluded that the rapid density change is the result of the fluid extraction and not the flow impediment of red blood cells caused by the increase in osmolarity.

Animals↗

Permeability studies on red cell membranes of dog, cat, and beef.

Water permeability coefficients of dog, cat, and beef red cell membranes have been measured under an osmotic pressure gradient. The measurements employed a rapid reaction stop flow apparatus with which cell shrinking was measured under a relative osmotic pressure gradient of 1.25 to 1.64 times the isosmolar concentration. For the dog red cell the osmotic permeability coefficient is 0.36 cm(4)/(sec, osmol). The water permeability coefficient for the dog red cell under a diffusion gradient was also measured (rate constant = 0.10/msec). The ratio between the two permeabilities was used to calculate an equivalent pore radius of 5.9 A. This value agrees welt with an equivalent pore radius of 6.2 A obtained from reflection coefficients of nonelectrolyte water-soluble molecules, and is consistent with data on the permeability of the dog red cell membrane to glucose. These data provide evidence supporting the existence of equivalent pores in single biological membranes.

Animals↗

A new proposal for the action of vasopressin, based on studies of a complex synthetic membrane.

The total osmotic flow of water across cell membranes generally exceeds diffusional flow measured with labeled water. The ratio of osmotic to diffusional flow has been widely used as a basis for the calculation of the radius of pores in the membrane, assuming Poiseuille flow of water through the pores. An important assumption underlying this calculation is that both osmotic and diffusional flow are rate-limited by the same barrier in the membrane. Studies employing a complex synthetic membrane show, however, that osmotic flow can be limited by one barrier (thin, dense barrier), and the rate of diffusion of isotopic water by a second (thick, porous) barrier in series with the first. Calculation of a pore radius is meaningless under these conditions, greatly overestimating the size of the pores determining osmotic flow. On the basis of these results, the estimation of pore radius in biological membranes is reassessed. It is proposed that vasopressin acts by greatly increasing the rate of diffusion of water across an outer barrier of the membrane, with little or no accompanying increase in pore size.

Acetates↗