Mice stressed daily by brief cold water immersions for 1, 8 or 14 days showed changes in immune system function which were dependent on the number of mice per cage, frequency of stress exposures and total number of stress exposures. Changed percentages of spleen B and CD4, but not of CD8 cells were determined when the mice were stressed either once or twice daily. With CD4 cells, increased percentages were seen after stress once daily but a decreased percentage was seen after stress twice daily. Furthermore, the Concanavalin A-stimulated spleen cell mitogenesis was decreased after 1 day of stress in mice stressed once daily as opposed to after 8 and 14 days of stress in mice stressed twice daily. After 14 days of stress, the lipopolysaccharide stimulated mitogenesis was increased if the mice were stressed once daily but decreased if the mice were stressed twice daily. With two mice per cage, we observed a decreased spleen cell mitogenesis after 14 days of stress. With four mice per cage, the spleen cell mitogenesis was decreased after 8 and 14 days of stress. If spleen cell populations from mice stressed twice daily for 8 days were depleted of macrophages and CD4 or CD8 cells, the effect of stress on the mitogenesis was removed from the CD8 cells. Spleen cells of mice stressed for 14 days showed a decreased mitogenesis when depleted of adherent cells and reconstituted with adherent cells from control mice. Furthermore, the adherent cells from these mice had decreased ability to support mitogenesis of adherent cell-depleted spleen cells from control mice as well as a decreased IL-1 production.