Mouse L-929 cells (a fibroblast-like line) were transferred from normal growth medium to one supplemented with 0.3 M sorbitol, doubling the normal external osmotic pressure. After a short lag phase and minimal cell death, the cells began to grow, and the growth rate reached that of controls after about one week. These chronically grown cells (S) have been compared to those of control cultures (C) with regard to general morphology, ability to reverse when returned to normal condition, water content, volume and selected metabolic parameters. S-cell cultures exhibited considerable heterogeneity but most contained vesicle-like cytoplasmic structures, sometimes in abundance. These structures do not appear to be completely bounded by membranes, but that is uncertain. S cells become larger and contain more water than C cells; however, the ratio of total water to total dry mass is indistinguishable from controls suggesting regulation at that level. S and C cells were found to be remarkably similar, on a per cell basis, with regard to their rate of respiration and the incorporation of glucose into metabolites and macromolecules. These results are interpreted in terms of current views on the composition and organization of the aqueous compartments of animal cells.