The proportions of bile acid (BA) species in the cytosol and nucleus of hepatocytes are different. The aim of the present study was to investigate the existence of selective access to hepatocyte nuclei of small organic molecules. Rat hepatocytes were incubated with 10-to-200 mM radiolabeled TCA, TUDCA, TCDCA or TDCA; nuclei were then isolated by lysis and differential centrifugation, and the radioactivity determined. Non-linear relationships between cellular BAs uptake by isolated hepatocytes and their nuclear content were found. When rat hepatocytes or suspensions of nuclei isolated from these cells were incubated with fluorescent BA derivatives CGamF, UDCGamF or CDCGamF, it was possible to detect these compounds in the nuclei by confocal microscopy. They were not homogenously distributed, but localized in DNA-poor regions. Measurement by flow cytometry revealed that the uptake of FITC was 3-to-4-fold reduced by conjugation with a BA moiety. Doxorubicin but scarcely rhodamine and FITC-dextrans (10kDa>40kDa=70kDa) were also incorporated into these organelles. The presence of ATP reduced nuclear load of these compounds. This effect was not modified by the presence of vanadate or cyclosporin A and was also observed when ATP was replaced by a non-hydrolyzable ATP analog. Moreover, UDCGamF efflux from nuclei previously loaded was stimulated by the presence of ATP and the absence of Ca²⁺ in the incubation media. In conclusion: in the hepatocyte nucleus there are ATP-sensitive mechanisms able to selectively restrict the access or enhance the efflux of different small-size compounds, which may behave as ligands for nuclear receptors or be potentially genotoxic.