TRANSPORT OF WATER IN PROXIMAL KIDNEY-TUBULES - FROM WHOLE TUBULES TOSINGLE CHANNELS - LENGTH AND SECTION OF THE SELECTIVITY FILTER OF AQUAPORIN-1

Proximal straight tubule (PST) were dissected from rabbit kidneys, hel d with crimping pipettes in a chamber bathed in a buffered mannitol is osmotic solution (MBS, 295 mOsm/kg). Tubule cell volume changes with t ime (dV/Adt) after steps in MBS osmolality (Delta C-s) were monitored on line with an inverted microscope, a TV camera and an image processo r. Reflection coefficients sigma and osmotic permeability coefficients , P-os, for several solutes were measured using two methods. Method I: a was calculated from the Delta C-s(iso) of impermeant and permeant s olutes at which (dV/Adt)(t-->0) = 0 (i.e., by a null point method). It is denoted as sigma(1).sigma(1) = 1.00 for mannitol (M), raffinose (R ), sucrose (S), glycerol (G), acetamide (A) and urea (U). With formami de (F), sigma(1.Formamide) = 0.62 +/- 0.05. These findings confirm our previous value of d(p) 4.5 Angstrom for the diameter of the selectivi ty filter of the basolateral PST cell membrane water channel AQP1. Met hod 2: PST were exposed for 20 s to MBS made hyperosmotic by addition of a Delta C-s of 35 mOsm/kg of R, S, M, G, A and U. Cells shrunk with in 500 ms of t = 0 to their osmometric volume and remained shrunk for the 20 s of the osmotic challenge. P-os was measured from the shrinkin g curves. P-os = 3000 +/- 25 mu m/s with R, S, M, G, A and U. Method 2 also allowed to calculate sigma denoted as sigma(2). sigma(2) = 1.00 for R, S, M, G, A and U. By contrast, the shrinking curve produced by a Delta C-s of 35 mOsm/kg F was 1/5th to 1/6th slower and smaller (i.e ., subosmometric) than that produced by a Delta C-s of 35 mOsm/kg R, S , M, G, A and U. Furthermore, with F cells did not remain shrunk but r ecovered their original volume within 3 s. P-os (measured with F) is d enoted as P-os, P-os* = 480 +/- 30 mu m/s. sigma(2.Formamide) = 0.16 +/- 0.01. Use of sigma(1), sigma(2) and P-os values in Hill's equatio ns for the bimodal theory of osmosis leads to n = 2-9. Where n is the number of water molecules single filling within the channel selectivit y filter, whose length must lie within 6 to 27 Angstrom, a value signi ficantly lower than our previous value calculated from the P-os/P-d r atio.