Hydrothermal treatment of anodized alumina membranes has been known for years and is believed to seal the pores by transforming aluminium oxide into lower density hydroxides. We demonstrate that, at least for 60 nm diameter pores grown from anodization in oxalic acid at 40 V, the hydrothermal treatment significantly shrinks but does not fully seal the nanopores. The pores shrink to a neck of less than 10 nm in diameter and 2–4 µm in length, in which the diffusion coefficient of ions is five orders of magnitude smaller than in the bulk. Because of a high electrolyte resistance through hydrothermally treated shrunken nanopores, they can be used for electrical sensing applications, as demonstrated using the example of DNA sensing. Hybridization of target DNA with a complementary ssDNA covalently immobilized inside the nanopores causes an increase in impedance by more than 50% while a noncomplementary ssDNA has no measurable effect.