The protozoan Giardia lamblia initiates infection when trophozoites emerge from a cyst in the hosts by the excystation process. Although this process is crucial to the initiation of infection by G. lamblia, little is known about its regulation. To study the possible involvement of calmodulin (CaM) in excystation we tested the effect of several CaM antagonists (TFP, W-7, and W-5) on this cellular function. Except for W-5 the rest of these compounds inhibited excystation. The protein kinase C inhibitor H-7 had no effect on excystation, suggesting that CaM antagonists acted by selectively inhibiting CaM. Furthermore, CaM was redistributed after the induction of excystation and there was an increase in its fluorescence and activity. These results suggest that a CaM-dependent process is involved in G. lamblia excystation.