A simple technique to efficiently dissociate primary auditory neurons from 5 day-old rat cochleas.

The aim of this work was to develop a simple and reproducible method of dissociation of cochlear spiral ganglion neurons in the rat. This technique, wich was developed in 5 day-old rat pups, was based on the use of a single enzyme, thermolysin. It is easy to set up and allows the collection of a large amount of neurons. These isolated neurons were kept in a definite, serum free culture medium up to 7 days. Neurons were characterized both by standard morphological criteria and by using a specific neuronal marker (anti-neurofilament 200 kD) after 2 h and 7 days in culture. Cell viability, assessed by fluorescent dyes indicated that all isolated cells were healthy even after 7 days in vitro. The dissociation and culture methods were found very satisfactory and can be easily adapted to any kind of experiment requiring isolated spiral ganglion neurons.