High yield extraction of pure spinal motor neurons, astrocytes and microglia from single embryo and adult mouse spinal cord

• Published in: Scientific reports 2015-11, Vol.5 (1), p.16763-16763, Article 16763
• Main Authors: Beaudet, Marie-Josée, Yang, Qiurui, Cadau, Sébastien, Blais, Mathieu, Bellenfant, Sabrina, Gros-Louis, François, Berthod, François
• Format: Article
• Language: English
• Subjects: 13/106; 13/109; 13/31; 13/51; 14; 14/19; 631/378/87; 631/80/304; 692/308/1426; 692/617/375/1917/1285; Amyotrophic lateral sclerosis; Animals; Astrocytes; Astrocytes - cytology; Astrocytes - metabolism; Biomarkers; Cell culture; Cell Separation; Cell Survival; Cells, Cultured; Digestion; Embryo, Mammalian - cytology; Embryo, Mammalian - metabolism; Embryos; Gene Expression; Humanities and Social Sciences; Mice; Mice, Transgenic; Microglia; Microglia - cytology; Microglia - metabolism; Motor neurons; Motor Neurons - cytology; Motor Neurons - metabolism; Motor task performance; multidisciplinary; Neurodegenerative diseases; Neuronal-glial interactions; Phenotype; Rodents; Science; Spinal cord; Spinal Cord - cytology; Spinal Cord - metabolism; Transgenic mice
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/srep16763

Extraction of mouse spinal motor neurons from transgenic mouse embryos recapitulating some aspects of neurodegenerative diseases like amyotrophic lateral sclerosis has met with limited success. Furthermore, extraction and long-term culture of adult mouse spinal motor neurons and glia remain also challenging. We present here a protocol designed to extract and purify high yields of motor neurons and glia from individual spinal cords collected on embryos and adult (5-month-old) normal or transgenic mice. This method is based on mild digestion of tissue followed by gradient density separation allowing to obtain two millions motor neurons over 92% pure from one E14.5 single embryo and more than 30,000 from an adult mouse. These cells can be cultured more than 14 days in vitro at a density of 100,000 cells/cm 2 to maintain optimal viability. Functional astrocytes and microglia and small gamma motor neurons can be purified at the same time. This protocol will be a powerful and reliable method to obtain motor neurons and glia to better understand mechanisms underlying spinal cord diseases.