Subcellular fractionation of brain tissue from small tissue explants

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, the brain tissue is built through the networking of neuronal, glial, and vascular cells in an intermingled meshwork of micrometer-sized structures. Subcellular fractionation protocols allow for the separation of specific compartments such as synapses (called “synaptosomes”), synaptic plasma membranes, and synaptic vesicles for their analysis at the molecular level. Most current protocols were established to process large amounts of tissue as required in previous experimental paradigms. Here, we provide a protocol to prepare synaptosomes from as little as 10 mg of tissue or a full fractionation to enrich crude synaptic vesicles and synaptic plasma membranes from 20 mg of tissue. This protocol will be useful to anyone aiming at addressing specific questions regarding local microcircuits in combination with connectomics and proteomics approaches.