Carbachol and histamine both stimulate calcium-dependent chloride secretion in the colonic epithelial cell line, T84. However, pretreatment of cell monolayers with carbachol blocks subsequent chloride secretion induced by thapsigargin but not the calcium elevation stimulated by this agent, whereas histamine pretreatment blocks neither thapsigargin-induced chloride secretion nor calcium elevation. To examine whether inositol phosphate metabolism might account for this difference, we measured levels of radiolabeled inositol phosphates: Ins(1,3,4)P3, Ins(1,4,5)P3, Ins(1,3,4,5)P4, Ins-(1,3,4,6)P4, Ins(3,4,5,6)P4, InsP5, and InsP6 after cell stimulation. Although both carbachol and histamine increase Ins (1,4,5)P3 at 5 s, there is a greater and more persistent increase in the levels of Ins(1,3,4)P3 and InsP4 at later time points after carbachol than histamine, which corresponded to the suppression of the chloride secretory response.