The cellular and molecular mechanisms involved in osteoblastic cell replication induced by strontium ranelate are presently under investigation. The calcium-sensing receptor is a suggested target but other potential mechanisms have not been investigated. Signaling pathways involved in strontium ranelate-induced replication were investigated in preosteoblastic MC3T3-E1 and pluripotent mesenchymal C3H10T1/2 cells. Strontium ranelate effects were compared with those of calcium chloride as Ca(2+). In MC3T3-E1 cells, strontium ranelate but not CaCl(2) dose-dependently increased cell number whereas similar effects were observed for both cations in C3H10T1/2 cells. Immunoblot analysis indicated that activation of ERK, PKC and PKD by strontium ranelate in MC3T3-E1 cells was delayed compared with CaCl(2). Indeed, onset of signaling by strontium ranelate was detected after one or several hours whereas CaCl(2) had a maximal effect already after 5 min exposure. In C3H10T1/2 cells, strontium ranelate induced two types of signaling, a rapid effect and a delayed response. In addition to activation of ERK, PKC and PKD, strontium ranelate and CaCl(2) also transiently activated p38 in C3H10T1/2 cells. Functional analysis with specific inhibitors indicated that cell replication induced by strontium ranelate involves a PKC/PKD pathway in MC3T3-E1 cells and p38 in C3H10T1/2 cells. In both cell types, inhibition of the ERK pathway decreased basal cell replication but not the strontium ranelate response. In conclusion, strontium ranelate increases the replication of cells of the osteoblastic lineage by two distinct cellular mechanisms. Strontium ranelate may directly interact with the CaSR and trigger mitogenic signals such as p38 in C3H10T1/2 cells. The delayed activation of several signaling pathways in both cell lines, however, suggests the release of an autocrine growth factor by strontium ranelate that represents another potential mechanism for inducing osteoblastic cell replication.