Dissection, Plating, and Maintenance of Dorsal Horn Neuron Cultures: Figure 1.

Abstract

INTRODUCTIONPrimary cultures of embryonic dorsal horn (DH) neurons can be prepared using mass cultures, microisland cultures, or macroisland cultures. In all cases, long-term survival of neurons is enhanced by the presence of astrocytes. After the feeder layer of astrocytes has become nearly or fully confluent, it is ready to receive freshly dissociated neurons. Embryonic dorsal root ganglion (DRG) neurons can also be added to the cultures if DRG/DH neuron cocultures are desired. These cultures are suitable for electrophysiological, molecular, and immunocytochemical studies. DH neurons, including both excitatory and inhibitory neurons, form synapses with themselves (autapses) and with each other. DRG neurons form synapses onto DH neurons. Finding synaptically connected pairs of neurons is most readily achieved when the cells are grown on microislands. This protocol describes the dissociation, plating, and maintenance of DH neuron cultures using a feeder layer of astrocytes.