The stromal interaction molecules STIM1 and STIM2 sense a decreasing Ca(2+) concentration in the lumen of the endoplasmic reticulum and activate Ca(2+) channels in the plasma membrane. In addition, at least 2 reports suggested that STIM1 may also interact with the inositol 1,4,5-trisphosphate (IP(3)) receptor. Using embryonic fibroblasts from Stim1(-/-), Stim2(-/-) and wild-type mice, we now tested the hypothesis that STIM1 and STIM2 would also regulate the IP(3) receptor. We investigated whether STIM1 or STIM2 would be the luminal Ca(2+) sensor that controls the loading dependence of the IP(3)-induced Ca(2+) release. Partial emptying of the stores in plasma-membrane permeabilized cells resulted in an increased EC(50) and a decreased Hill coefficient for IP(3)-induced Ca(2+) release. This effect occurred both in the presence and absence of STIM proteins, indicating that these proteins were not the luminal Ca(2+) sensor for the IP(3) receptor. Although Stim1(-/-) cells displayed a normal IP(3)-receptor function, agonist-induced Ca(2+) release was reduced. This finding suggests that the presence of STIM1 is required for proper agonist-induced Ca(2+) signaling. Our data do not provide experimental evidence for the suggestion that STIM proteins would directly control the function of the IP(3) receptor.