The effect of anisotonic solutions on the enhancement of the transport of hydrophilic model compounds across monolayers of Caco-2 and HT-29.cl19A intestinal epithelial cells was studied. In filter-grown monolayers of the highly differentiated villus-like Caco-2 cell line, a profound and dose-dependent drop in the transepithelial electrical resistance was found after apical treatment with a 30 or a 50% hypotonic solution (200 and 150 mOsmol, respectively). This drop was not observed after basolateral and two-sided application of a 50% hypotonic solution. During apical hypotonic treatment a 12- and 8-fold increase also was observed in transepithelial transport of two hydrophilic model compounds, i.e., fluorescein-Na and fluorescein-isothiocyanate-labeled dextran, MW 4000, respectively. Through confocal laser scanning microscopy, it was revealed that this enhanced transport was predominantly via the paracellular route. Moreover, morphological changes in the cell layers indicating cell swelling were observed after apical hypotonic, but not after basolateral or bilateral treatment, probably resulting from an incomplete regulatory volume decrease response. This swelling, and slight lateral retraction of the cells, allowed the hydrophilic compounds to pass between the cells. The effects of hypotonic challenge also were studied in monolayers of the more crypt cell-like HT-29.cl19A cell line. After apical hypotonic shock, these cells showed no effect on transepithelial electrical resistance, whereas an increase was observed after basolateral and bilateral treatment. Hypotonic shock failed to increase the transport of the hydrophilic model compounds in this cell line.(ABSTRACT TRUNCATED AT 250 WORDS)