Increasing the osmolarity of the culture medium enhances the response of peanut agglutinin (PNA)-negative thymocytes to stimulation by 12-O-tetradecanoylphorbol-13-acetate (TPA), interleukin 1 (IL-1) and interleukin 2 (IL-2) in the presence of phytohemagglutinin (PHA). The effect was attained by the addition to the medium of salts such as NaCl and KCl or by addition of nonionized compounds such as sucrose and fucose. The enhanced response was monitored by determination of [3H]thymidine incorporation, IL-2 production, and blasts formation. The potentiating effect of hypertonic medium on PNA-negative thymocytes treated with PHA and TPA was most pronounced at suboptimal concentrations of PHA. Hypertonic medium did not enhance the response of thymocytes treated with TPA and supraoptimal concentrations of PHA. Increasing the osmolarity of the medium 44 hr after initiation of culture did not enhance [3H]thymidine incorporation in thymocytes that were pulsed between 52 and 72 hr. The enhancing effect of increased osmolarity in mitogenic stimulation of thymocytes may be related to osmotic activation of the Na+ H+ antiport.
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