Escherichia coli cells were used to study the mechanism of penetration of local anesthetics and the relationship between permeation and functional properties. We show that both the neutral and the protonated form of dibucaine can be accumulated in the cells. Accumulation of the protonated form occurs in response to a transmembrane electrical potential (negative inside) and results in high trapped concentrations (70 mM). Accumulation can lead to an alkalinization of the internal pH. Low concentrations of dibucaine stimulate the respiration, increase the transmembrane electrical potential and raise the accumulation of solutes. Inhibition of these functions occurs at higher concentrations of the drug. Furthermore, the drug concentration required to inhibit these functions is smaller at alkaline external pH than at acidic external pH, suggesting that the inhibition is mainly due to the neutral form of the anesthetics. Other hydrophobic amines also stimulate and inhibit different membrane functions, their efficiency being correlated to their lipophilicity.