Osmotic water permeability of small intestinal brush-border membranes

Abstract

A stopped-flow nephelometric technique was used to examine osmotic water flow across small intestinal brush-border membranes. Brush-border membrane vesicles (BBMV) were prepared from rat small intestine by calcium precipitation. Scattered 500 nm light intensity at 90° to incident was a linear function of the number of vesicles in suspension, and of the reciprocal of the suspending medium osmolality. When BBMV were mixed with hyperosmotic mannitol solutions there was a rapid increase in the intensity of scattered light that could be fit to a single exponential function. The rate constant for vesicle shrinking varied with temperature and the size of the imposed osmotic gradient. At 25°C and an initial osmotic gradient of 50 mOsm, the rate constant was 1.43±0.044 sec⁻¹. An Arrhenius plot of the temperature dependence of vesicle shrinking showed a break at about 25°C with an activation energy of 9.75±1.04 kcal/mole from 11 to 25°C and 17.2±0.55 kcal/mole from 25 to 37°C. The pore-forming antibiotic gramicidin increased the rate of osmotically driven water efflux and decreased the activation energy of the process to 4.51±0.25 kcal/mole. Gramicidin also increased the sodium permeability of these membranes as measured by the rate of vesicle reswelling in hyperosmotic NaSCN medium. Gramicidin had no effect on mannitol permeability. Assuming spherical vesicles of 0.1 μm radius, an osmotic permeability coefficient of 1.2×10⁻³ cm/sec can be estimated for the native brush-border membranes at 25°C. These fesults are consistent with the solubility-diffusion model for water flow across small intestinal BBMV but are inconsistent with the existence there of large aqueous pores.