Lys-Bradykinin (BK), a mitogen for human foreskin fibroblasts (HSWP cells) (Owen, N. E., and Villereal, M. L. (1983) Cell 32, 979-985), elicits a rapid, transient elevation of intracellular free Ca2+ concentration ([Ca2+]i) in these cells. We have used image analysis of fura-2-loaded HSWP cells to examine the BK-induced [Ca2+]i changes in individual cells. BK-stimulated Ca2+ entry and release of intracellular Ca2+ stores can be distinguished by stimulating cells in the presence or absence of extracellular Ca2+, or by inhibiting Ca2+ entry with 5 mM NiCl2. BK-sensitive intracellular Ca2+ stores can be depleted by exposure of the cells to BK in Ca(2+)-free medium; refilling of the stores requires extracellular Ca2+. A component of BK-stimulated Ca2+ entry persists after removal of agonist, but inactivates with a t1/2 of approximately 5 min. Although previous studies have attributed the Ca2+ entry which persists after agonist removal to a "capacitative Ca2+ entry" pathway activated by the depletion of the intracellular Ca2+ stores, we find that a large component of this BK-stimulated Ca2+ entry is not due to capacitative Ca2+ entry since (1) ionomycin can deplete the BK-sensitive intracellular Ca2+ stores without appreciably stimulating Ca2+ entry and without inhibiting the BK-stimulated Ca2+ entry and (2) this Ca2+ entry pathway inactivates at a time when the Ca2+ pools are still empty and a capacitance entry pathway should still be open. On the other hand, refilling of the intracellular Ca2+ stores can occur after the noncapacitative Ca2+ entry component has inactivated or when it is inhibited by Ni2+; in these cases refilling occurs without a detectable elevation of [Ca2+]i suggesting that refilling of internal Ca2+ pools might occur by a capacitative route.