An efficient and cost-effective method of generating postnatal (P2-5) mouse primary hippocampal neuronal cultures

Primary culture of postnatal central neurons is a widely used methodology for applications such as the investigation of neuronal development, protein trafficking/distribution and cellular signalling. However, successful production and maintenance of such cultures, particularly from postnatal animals, can be challenging. In attempting to surmount these difficulties, several disparate culturing methodologies have been developed. Such methodologies are centred on the identification and optimisation of critical steps and, as such, the protocols and reagents utilised can differ quite markedly from protocol to protocol, often with the suggestion that the use of a (usually expensive) proprietary reagent(s), lengthy substrate preparation and/or cell isolation techniques is/are necessary for successful culture preparation.

Herein, we present a simple and inexpensive protocol for the preparation of primary hippocampal neurons from postnatal (2-5 day old) mice, which remain viable for experimental use for over one month.

Neurons cultured using this method follow well established developmental norms and display typical responses to standard physiological stimuli such as depolarisation and certain pharmacological agents.

By using a novel trituration technique, simplified methodology and non-proprietary reagents, we have developed a reliable protocol that enables the cost effective and efficient production of high quality postnatal mouse hippocampal cultures. This method, if required, can also be utilised to prepare neurons both from other regions of the brain as well as from other species such as rat.