We conducted a comparative study of NK1.1+ cells in spleen and bone marrow and the effects of administration of M-CSF on them. Administration of M-CSF to mice increased the number of NK1.1+ cells in spleen but not in bone marrow. The NK1.1+ cells in spleen (Spl-NK1.1) and bone marrow (BM-NK1.1) were purified by magnetic cell sorter. Their cell surface markers and functions were then examined. The percentage of Mac-1 antigen-positive cells (and F4/80 antigen-positive cells) was higher among BM-NK1.1 than Spl-NK1.1. Moreover, the administration of M-CSF increased the number of Mac-1 and F4/80 antigen-positive cells in both Spl-Nk1.1 and BM-NK1.1. The functions (cytolytic activity and IFN-gamma production) of Spl-NK1.1 and BM-NK1.1 were the same and were enhanced by the administration of M-CSF. But Spl-NK1.1 produced more IFN-gamma than BM-NK1.1 when M-CSF was administered. BM-NK1.1 showed a greater proliferative response to IL-2 than Spl-NK1.1. Administration of M-CSF augmented this response. BM-NK1.1 proliferated in response to IL-4 and IL-15, but Spl-NK1.1 responded only slightly. However, administration of M-CSF stimulated Spl-NK1.1 to respond to these cytokines. Both Spl-NK1.1 and BM-NK1.1 showed only a weak response to M-CSF in vitro. But the expression of c-fms antigen (M-CSFR) increased after the M-CSF injections in vivo. These results suggested that there are phenotypical and functional differences between Spl-NK1.1 and BM-NK1.1. The administration of M-CSF led to an accumulation of NK1.1+ cells which were mobilized from bone marrow in spleen.