On impermeable substrata MDCK cells, a cell line derived from normal dog kidney, forms a confluent monolayer that is studded with numerous hemicysts. Previous studies with this cell line suggest that thes hemicysts develop as a result of active fluid accumulation between cell sheet and substratum. However, the formation of hemicysts as a multifocal phenomenon is still unexplained. The results presented here show that the hemicysts are not only expressions of active transport of solutes and water, but also of cell-substratum interaction. The increase in number and size of the hemicyst produced by dbcAMP may be explained by a decrease in the adhesive strength to substrata produced by this compound. Moreover, when the strength of the cell-substratum adhesion was increased the number of hemicysts was reduced or abolished. On the contrary, when this strength was reduced, larger hemicysts occurred, covering practically all the area available for growth. Results from cinematographic time lapse studies, showing that 90% of the area of the monolayer is able to produce hemicysts, also suggest that hemicyst formation as a multifocal phenomenon is more an expression of local variations in cell-substratum interaction than of regional changes in transepithelial active transport.