Purification of Synaptosome Populations Using Fluorescence-Activated Synaptosome Sorting.

Abstract

For several decades, neurobiologists have used subcellular fractionation methods to analyze the molecular structure and some functional features of the cells in the central nervous system. Indeed, brain tissue contains a complex intermingled network of neuronal, glial, and vascular cells. To reduce this complexity biochemists have optimized fractionation protocols that enrich in specific compartments such as synapses (called "synaptosomes") and synaptic vesicles, for example. However, recently, these approaches suffered from a lack of specificity and purity. In a recent effort, we extended the conventional synaptosome preparation to purify fluorescent synaptosomes on a cell sorter. We could prove that our method allows for the steep enrichment in fluorescent excitatory VGLUT1venus synaptosomes containing the presynaptic element and the tip of the post-synaptic element and a strong depletion in neuronal and glial contaminants. Here, we propose a detailed procedure for the implementation of Fluorescence Activated Synaptosome Sorting.