Hydraphiles are synthetic ion channels that use crown ethers as entry portals and that span phospholipid bilayer membranes. Proton and sodium cation transport by these compounds has been demonstrated in liposomes and planar bilayers. In the present work, whole cell patch clamp experiments show that hydraphiles integrate into the membranes of human embryonic kidney (HEK 293) cells and significantly increase membrane conductance. The altered membrane permeability is reversible, and the cells under study remain vital during the experiment. Control compounds that are too short (C8-benzyl channel) to span the bilayer or are inactive owing to a deficiency in the central relay do not induce similar conductance increases. Control experiments confirm that the inactive channel analogues do not show nonspecific effects such as activation of native channels. These studies show that the combination of structural features that have been designed into the hydraphiles afford true, albeit simple, channel function in live cells.