Accumulating evidence suggests that histamine synthesis induced in several types of tumor tissues should modulate tumor immunity. We found that a transient histamine synthesis was induced in CD11b+Gr-1+splenocytes derived from BALB/c mice transplanted with a syngeneic colon carcinoma, CT-26, when they were co-cultured with CT-26 cells. Significant levels of IFN-γ were produced under this co-culture condition. We explored the modulatory roles of histamine on IFN-γ production and found that several histamine receptor antagonists, such as pyrilamine, diphenhydramine, JNJ7777120, and thioperamide, could significantly suppress IFN-γ production. However, suppression of IFN-γ production by these antagonists was also found when splenocytes were derived from the Hdc-/- BALB/c mice. Suppressive effects of these antagonists were found on IFN-γ production induced by concanavalin A or the combination of an anti-CD3 antibody and an anti-CD28 antibody in a histamine-independent manner. Murine splenocytes were found to express H1 and H2 receptors, but not H3 and H4 receptors. IFN-γ production in the Hh1r-/- splenocytes induced by the combination of an anti-CD3 antibody and an anti-CD28 antibody was significantly suppressed by these antagonists. These findings suggest that pyrilamine, diphenhydramine, JNJ7777120, and thioperamide could suppress IFN-γ production in activated splenocytes in histamine-independent manner.