CONCENTRATION POLARIZATION OF FLUORESCENT DYES IN RAT DESCENDING COLONIC CRYPTS - EVIDENCE OF CRYPT FLUID ABSORPTION

1. Using confocal microscopy, the rate of fluid absorption into isolat ed perifused descending rat colonic crypt lumens is estimated from the concentration polarization and distribution of fluorescein sulphonate (FS) and fluorescein isothiocyanate-dextran (FITC-dextran; molecular weight, 10 000) within the crypt lumens and pericryptal fluid. 2. The probe dyes enter the crypt via the luminal. opening, are concentrated in the lumen, then escape into the pericryptal space via the paracellu lar spaces spanning the crypt wall. 3. FITC-dextran is maximally accum ulated at a luminal depth of 60 mu m to 5 times the concentration at t he crypt opening (p < 0.001) and penetrates 150-200 mu m along the lum en. FX is maximally accumulated within crypt lumen close to the openin g. At crypt luminal depths 10-60 mu m from the opening FS is accumulat ed by a factor of 1.5-2.0 above that found in HgCl2-treated tissue (p < 0.001). 4. Dye enters the crypt lumen slowly from the basal side, bu t from this side does not accumulate above the bathing solution concen tration. 5. HgCl2 (20 mu M) or theophylline (10 mM) completely inhibit concentrative accumulation of FITC-dextran and FS within the crypts a nd pericryptal space (p < 0.001). 6. Computer simulation of the dye up take indicates that the rate of water flow into the crypt luminal open ing is 1 x 10(-3) cm s(-1) which is equivalent to 15 mu l (cm mucosa)( -2) h(-1). Approximately 75% of the fluid entering the crypt is absorb ed across the proximal 50 mu m of crypt wall as a consequence of the l arge osmotic pressure gradient between the pericryptal and crypt lumin al solutions. A pericryptal diffusion barrier with lower permeability than that across the crypt wall is required to simulate dye accumulati on in the pericryptal space. Differences between FITC-dextran and FS a ccumulation are explained by the lower diffusion coefficient within th e crypt lumen, and lower crypt wall permeability of FITC-dextran.