In the present study apolipoprotein-mediated free cholesterol (FC) efflux was studied in J774 macrophages having normal cholesterol levels using an experimental design in which efflux occurs in the absence of contributions from cholesteryl ester hydrolysis. The results show that cAMP induces both saturable apolipoprotein (apo) A-I-mediated FC efflux and saturable apo A-I cell-surface binding, suggesting a link between these processes. However, the EC₅₀ for efflux was 5–7-fold lower than the K_d for binding in both control and cAMP-stimulated cells. This dissociation between apo A-I binding and FC efflux was also seen in cells treated for 1 h with probucol which completely blocked FC efflux without affecting apo A-I specific binding. Thus, cAMP-stimulated FC efflux involves probucol-sensitive processes distinct from apo A-I binding to its putative cell surface receptor. FC efflux was also dramatically stimulated in elicited mouse peritoneal macrophages, suggesting that cAMP-regulated apolipoprotein-mediated FC efflux may be important in cholesterol homeostasis in normal macrophages. The presence of a cAMP-inducible cell protein that interacts with lipid-free apo A-I was investigated by chemical cross-linking of ¹²⁵I-apo A-I with J774 cell surface proteins which revealed a M_r 200 kDa component when the cells were treated with cAMP.