Validation of a chamber that allows measurement of both tissue uptake rates and unstirred layer thicknesses in the intestine under conditions of controlled stirring

It is now apparent that the study of transport mechanisms in the intestine must take cognizance of the presence of a significant diffusion barrier overlying the villus surface. This paper describes a new incubation chamber that allows measurement of both unstirred layer thicknesses and tissue uptake rates under identical conditions where the rate of stirring can be reproducibly and systematically varied. At a stirring rate of 500 rpm, the mean thickness of the diffusion barrier in rabbit jejunum was approximately 150 μm. Use of a marker of adherent mucosal fluid volume was shown to be of critical importance in obtaining reliable transport data, particularly when studying poorly permeant solutes or solutes solubilized in bile acid micelles. The volume of the adherent mucosal fluid varied markedly depending on such experimental variables as the presence of intestinal mucus or bile acids in the incubation solution. At least 3 to 6 min were required to fully equilibrate the unstirred layers with a marker compound such as dextran or inulin that was used to calculate the adherent mucosal fluid volume. On the other hand, incubations of more than 10 min led to loss of solute molecules into the serosal compartment. Thus, in order to obtain reliable uptake rates, measurements had to be made at 6 to 10 min of incubation in this chamber; shorter or longer incubation times, respectively, led to over or underestimates of flux rates.