Zebrafish (Danio rerio) is increasingly used to assess the pharmacological activity and toxicity of compounds. The spatiotemporal distribution of seven fluorescent alkyne compounds was examined during 48 h after immersion (10 µM) or microinjection (2 mg/kg) in the pericardial cavity (PC), intraperitoneally (IP) and yolk sac (IY) of 3 dpf zebrafish eleuthero-embryos. By modelling the fluorescence of whole-body contours present in fluorescence images, the main pharmacokinetic (PK) parameter values of the compounds were determined. It was demonstrated that especially in case of short incubations (1-3 h) immersion can result in limited intrabody exposure to compounds. In this case, PC and IP microinjections represent excellent alternatives. Significantly, IY microinjections did not result in a suitable intrabody distribution of the compounds. Performing a QSPkR (quantitative structure-pharmacokinetic relationship) analysis, LogD was identified as the only molecular descriptor that explains the final uptake of the selected compounds. It was also shown that combined administration of compounds (immersion and microinjection) provides a more stable intrabody exposure, at least in case of a prolonged immersion and compounds with LogD value > 1. These results will help reduce the risk of false negative results and can offer an invaluable input for future translational research and safety assessment applications.
The blood-brain barrier (BBB) is an efficient barrier for molecules and drugs. Multicellular 3D spheroids display reproducible BBB features and functions. The spheroids used here were composed of six brain cell types: Astrocytes, pericytes, endothelial cells, microglia cells, oligodendrocytes, and neurons. They form an in vitro BBB that regulates the transport of compounds into the spheroid. The penetration of fluorescent ultrasmall gold nanoparticles (core diameter 2 nm; hydrodynamic diameter 3-4 nm) across the BBB was studied as a function of time by confocal laser scanning microscopy, with the dissolved fluorescent dye (FAM-alkyne) as a control. The nanoparticles readily entered the interior of the spheroid, whereas the dissolved dye alone did not penetrate the BBB. We present a model that is based on a time-dependent opening of the BBB for nanoparticles, followed by a rapid diffusion into the center of the spheroid. After the spheroids underwent hypoxia (0.1% O2; 24 h), the BBB was more permeable, permitting the uptake of more nanoparticles and also of dissolved dye molecules. Together with our previous observations that such nanoparticles can easily enter cells and even the cell nucleus, these data provide evidence that ultrasmall nanoparticle can cross the blood brain barrier.
