Vol. 295, Issue 1, 367-372, October 2000

Cellular Uptake and Efficacy of Antisense Oligonucleotides against RNAs of Two Na⁺ Channel Isoforms¹

Hans Keller, Bernhard Schu, Reinhardt Rüdel and Heinrich Brinkmeier

Department of General Physiology, University of Ulm, Ulm, Germany

The effects of 15-mer phosphorothioate antisense oligodeoxynucleotides (aODNs) specifically designed against the RNAs of either of two closely related Na⁺ channel isoforms, hSkM1 or hH1, were tested in human myotubes. Fluorescence (3'-fluorescein isothiocyanate) labeling showed that mere incubation of cultures with aODNs did not result in aODN uptake, but liposome-mediated transfer was successful and resulted in cytoplasmic and nuclear localization of ODNs. Intracellular fluorescence was stable for at least 3 days. At 5 µM, the hH1-specific aODN was effective in suppressing ion channel function, but the hSkM1-specific aODN was not. Reverse transcription-polymerase chain reaction gave corresponding results on the mRNA level. However, in HEK-293 cells stably expressing hSkM1, the same hSkM1-specific aODN was able to reduce Na⁺ currents (2.4 ± 0.5 nA, n = 11; controls: 6.5 ± 1.0 nA, n = 14). We conclude that cellular uptake of aODNs and intracellular access to the RNA target are necessary, but not always sufficient conditions for an effective block of mRNA translation in intact cells.