WATER HANDLING IN CACO-2 CELLS - EFFECTS OF ACIDIFICATION OF THE MEDIUM

Caco-2 cells were cultured on permeable supports. At confluence the minute-by-minute net water movement (J(w)) was automatically recorded. Simultaneously, unidirectional [C-14]mannitol, Na-22+, and/or Cl-36 fluxes and transepithelial resistances were measured. The water and mannitol permeabilities went progressively down between 9 and 16 days after seeding and then stabilized. In this last condition the hydrostatic permeability coefficient (P(hydr)) was 2.67 +/- 0.31 cm s-1 while the osmotic permeability coefficient (P(osm)) was 0.001 7 +/- 0.0004 cm s-1. P(hydr) but not P(osm) was dependent on the temperature and on the presence of Na+ in the medium. A net secretory J(w) was observed 16 days after seeding, in the absence of any osmotic, hydrostatic or chemical gradient. This secretory J(w) was associated with net Cl- (1.43 +/- 0.43 muequiv h-1 cm-2) and Na+ (1.05 +/- 0.35 muequiv h-1 cm-2) secretions. Amiloride reduced, in open-circuit conditions, both Na+ and Cl- apical to basal fluxes, thus enhancing the net Na+ and Cl- exit. Acidification of the medium (pH 6.2) reversibly increased water and mannitol permeabilities in 10-day-old cultures. In 16-day-old cultures the same shift in medium pH did not change mannitol permeability, while stimulating water secretion. These results, obtained in the absence of supracellular structures (villae, crypts) and subepithelial components (muscular, vascular and conjunctive tissues) indicate that paracellular and transport-associated water pathways are sensitive to changes in the pH of the medium in Caco-2 cell layers.