Crossing of BBB using gold nanoparticles (AuNPS)

Delivery of pharmacologic agents to neurons in the brain across the blood brain barrier (BBB) remains a challenging task for neuroscientists. Because of their unique properties, nanoparticles have generated a great deal of interest for their potential use in therapeutics drug delivery. We investigated the use of colloidal gold (AuNP) in this context based on its characteristic stability as a metal nanoparticle. We explored the neuronal uptake of AuNPs both in vitro using SH-SY5Y human neuroblastoma cells and in vivo using direct brain injection in mice. AuNPs have a diameter of approximately 2 nm and an absorption spectrum with a peak at 350 nm. Due to their characteristic quantum size effect, the nanogold emission shifts from green to red upon forming clusters of approximately 200 nm in diameter. The clustered particles have a broad emission ranging from 400 to 600 nm. For the in vitro studies, we incubated SH-SY5Y cells with 1x105 AuNPs for 20 min, 1, 2 and 4 hrs. AuNP internalization was observed using a Leica microscope workstation B6000 equipped with an in vivo timelapse analysis capability. We confirmed our results using transmission electron microscopy (TEM). For in vivo, studies we injected a physiological solution containing 1x106 AuNPs into the carotid arteries of anesthetized mice (n=3). The physiological solution alone was injected into control mice for comparison. Our in vitro data demonstrated that gold-particles passed the cell membrane, clustered in the cytoplasm and induced apoptosis after 4 hrs of treatment. In vivo, a strong particle-associated florescence was observed in the cauda putamen region (CPu) 90 min after injection. Fluorescence was evenly distributed in the neuronal cells but was not observed in glial cells, as demonstrated by GFAP, Iba1 and NeuN immunoistochemistry. The cytotoxic effects of AuNPs were investigated both in vitro and in vivo using caspase-3 immunoreactivity and TUNEL assays for apoptosis. Our data suggest that AuNPs cross the BBB and have useful imaging properties that rely on clustering in cells, which changes their emission spectrum from green to red. Specific properties of these nanoparticles may offer new advantages as potential pharmaco-delivery tools in treatment of neurological diseases that may benefit from specific neuronal cell targeting