ANALYSIS OF THE STEADY-STATE DYNAMICS OF ORGANELLE MOTION IN CULTURED NEURITES - PUTATIVE INDICATOR OF NEUROTOXIC EFFECT

1. The objective of this study was to develop a physiologically based method to evaluate the neurotoxic potential of drug candidates in vitro. Rat embryo midbrain cells were grown in micromass culture, and the movement of mitochondria labelled with the fluorescent dye rhodamine 123 was quantified in fasciculated neurites, using a laser cytometer. 2. The rhodamine 123 signal in a defined region of fascicle was quantified and photobleached with the laser. A series of post-photobleach scans revealed the movement of fluorescent-labelled mitochondria into the bleached region from adjacent unbleached regions. Recovery of fluorescence is a measure of the size of the mobile pool of mitochondria relative to the total (moving plus stationary) pool. 3. The steady-state levels of fluorescence recovery was dependent on intracellular calcium and magnesium concentrations, energy status (ATP), and microtubule integrity (post taxol or vinblastine treatment). 4. This technique may be a useful indicator of neurotoxic effect.