Acute isolation of neurons from the mature mammalian central nervous system.

The acute dissociation procedure provides a simple means of isolating neurons from the mature mammalian central nervous system. The method was primarily devised to isolate neurons for patch-clamp electrophysiology. It may also prove useful for single-cell PCR, immunocytochemistry, sorting of fluorescently labeled cells, or long-term tissue culture of mature neurons. Dissociation is brought about by a combination of proteolysis and an ionic environment that encourages breakdown of the tissue. The method allows the isolation of neurons free of glial ensheathments in as little as 45 min after the sacrifice of the animal. Neurons so isolated lose fine dendritic branches, although the structure proximal to the cell body is often maintained, allowing identification of the morphological type of the neuron. The preparation has the following advantages: (1) the neurons are fully differentiated; (2) the cells are electronically compact, which improves the fidelity of the voltage clamp; (3) the cells are removed from the influence of surrounding cells; and (4) neurons can be isolated from small, circumscribed loci within the adult central nervous system.