Phagocytic microglia during delayed neuronal loss in the facial nucleus of the rat: time course of the neuronofugal migration of brain macrophages.

The injection of Fluoro-Gold (FG) into the whisker pad of rats yields a stable fluorescent labeling of the motoneurons in the lateral facial subnucleus. Following resection of 8-10 mm of the facial nerve, the microglia phagocytose the FG-preloaded neurons and assume the label. Employing this vital labeling of microglia in situ we studied the fate of same after completion of phagocytic activity. Starting at 56 days post resection (DPR) the FG-labeled microglia spread out from the lateral facial subdivision and invaded the entire facial nucleus. The quantitative analysis of this redistribution of the fluorescent marker revealed a prolonged increase in the number of labeled microglia strictly proportional to the delayed loss of neurons. The differentiation between microglia and shrunken neurons was performed with the new method of immunoquenching: the staining of vibratome sections with anti-rat neuron-specific enolase (NSE) combined with an ABC-HRP kit and DAB as detector totally extinguished (quenched) all fluorescence from the pre-labeled facial motoneurons. The fluorescent microglia were additionally stained with GSA I-B4 and OX-42, which should completely quench all fluorescence in the section. However, a few small round cells, always closely opposed to neuronal perikarya, still fluoresced. These NSE-negative, GSA I-B4 and OX-42 negative, but fluorescent cells may represent a new, immunologically uncharacterized microglial cell type, that participates in neuronophagia.