An Easy-to-Implement Protocol for Preparing Postnatal Ventral Mesencephalic Cultures
- 1Department of Chemical Engineering and Biotechnology, University of Cambridge, United Kingdom
- 2Department of Psychiatry, Columbia University, United States
Postnatally-derived cultures of ventral mesencephalic neurons offer several crucial advantages over embryonic ventral mesencephalic cultures, including a higher content of TH-positive cells and the ability to derive cells from the substantia nigra, which contains the neurons most vulnerable to Parkinson’s disease. On the other hand, these cultures are more challenging to produce consistently. Here, we provide an easy-to-implement protocol for culturing postnatal ventral mesencephalic cells from the substantia nigra (SN) and the ventral tegmental area (VTA) using commercially available media, dishes, and general lab equipment, avoiding extensive material and equipment purchases. The protocol can be completed in about 5 hours and provides ventral midbrain neuron (VM) cultures on cortex glia feeder layers in three weeks’ time. The protocol uses an optimized protease digestion, tissue storage in Hibernate A during dissection and purification of neurons on an OptiPrep density gradient.
Keywords: Dopaminergic Neurons, postnatal neuron cultures, Substantia Nigra, Ventral Tegmental Area, tyrosine hydroxylase, postnatal neurons, protocol, Parkinson’s disease
Received: 26 Oct 2017;
Accepted: 02 Feb 2018.
Edited by:Pier Giorgio Mastroberardino, Erasmus University Rotterdam, Netherlands
Reviewed by:Louis-Eric Trudeau, Université de Montréal, Canada
Ezia Guatteo, Parthenope University of Naples, Italy
Copyright: © 2018 Lautenschläger, Mosharov, Kanter, Sulzer and Kaminski Schierle1. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Dr. Janin Lautenschläger, University of Cambridge, Department of Chemical Engineering and Biotechnology, West Cambridge Site, Philippa Fawcett Drive, Cambridge, CB3 0AS, United Kingdom, firstname.lastname@example.org