Although P-glycoprotein (P-gp) is highly expressed in both intestinal epithelial cells and endothelial cells of brain capillaries, and functions as an efflux transporter in both organs, the impact of P-gp on intestinal absorption and brain uptake of drugs is quantitatively very different. The effect of P-gp on drug absorption is not quantitatively as important as suggested. Many drugs are good human P-gp substrates and yet exhibit reasonable oral bioavailability. In contrast, P-gp plays a quantitatively very important role in blocking the brain uptake of P-gp substrates. This review provides an overview of the role of P-gp in drug absorption and brain uptake, and explores possible factors that may explain the quantitative differences in the impact of P-gp on drug absorption and brain uptake.