Development and inheritance of osmotic tolerance in a line of spontaneously transformed BALB/3T3 cells

An in vitro line of transformed BALB/3T3 mouse fibroblasts was exposed to a 100 mM increase in the NaCl concentration of its growth medium. The rate of growth, as measured by the incorporation of tritiated thymidine, decreased almost 100-fold during the first 24 h of exposure to the hyperosmotic medium and then increased approximately 10-fold during the second 24 h of exposure. Cell counts of cultures passaged in medium with excess NaCl revealed a gradual increase in growth rate over a period of several weeks. The early kinetics by which salt tolerance developed in this cell line indicated a process of physiological adaptation rather than selection of preexisting variants in the control population. Clonally derived populations exposed to excess NaCl all showed a similar response. Cultures which tolerated a 100 mM increase in NaCl also grew well in medium containing 200 mM sucrose, indicating that their tolerance was not specific for NaCl. Although the initial response of cultures exposed to excess NaCl appeared to be one of physiological adaptation, tolerance for salt became hereditary after continued passage in hyperosmotic medium. Cultures that were returned to control conditions after prolonged exposure to excess NaCl inherited a high level of tolerance for salt which persisted for several hundred generations without selection.