The protozoan parasite Toxoplasma gondii infects its host cells through an active mechanism. In this work, we obtained evidence that host cells also play a fundamental role during the infection process. We found that previous incubation of the host cells, but not the parasites, with Dynasore, a small molecule that inhibits dynamin GTPase activity, markedly reduced the penetration of T. gondii tachyzoites into LLC-MK2 cells. In contrast, parasite adhesion to the host cell surface increased, as observed both by light and electron microscopy. Intriguingly, the few parasites internalized by Dynasore-treated cells remained in vacuoles located at the periphery of the cell, in contrast to the perinuclear localization seen in the control.