Bidirectional transport of molecules between nucleus and cytoplasm through the nuclear pore complexes (NPCs) spanning the nuclear envelope plays a fundamental role in cell function and metabolism. Nuclear import of macromolecules is a two-step process involving initial recognition of targeting signals, docking to the pore and energy-driven translocation. ATP depletion inhibits the translocation step. The mechanism of translocation itself and the conformational changes of the NPC components that occur during macromolecular transport, are still unclear. The present study investigates the effect of ATP on nuclear pore conformation in isolated nuclear envelopes from Xenopus laevis oocytes using the atomic force microscope. All experiments were conducted in a saline solution mimicking the cytosol using unfixed nuclear envelopes. ATP (1 mM) was added during the scanning procedure and the resultant conformational changes of the NPCs were directly monitored. Images of the same nuclear pores recorded before and during ATP exposure revealed dramatic conformational changes of NPCs subsequent to the addition of ATP. The height of the pores protruding from the cytoplasmic surface of the nuclear envelope visibly increased while the diameter of the pore opening decreased. The observed changes occurred within minutes and were transient. The slow-hydrolyzing ATP analogue, ATP-gamma-S, in equimolar concentrations did not exert any effects. The ATP-induced shape change could represent a nuclear pore "contraction."