Generating neurons from stem cells

• Published in: Methods in molecular biology (Clifton, N.J.) Vol. 438; p. 31
• Main Authors: Androutsellis-Theotokis, Andreas, Murase, Sachiko, Boyd, Justin D, Park, Deric M, Hoeppner, Daniel J, Ravin, Rea, McKay, Ronald D G
• Format: Journal Article
• Language: English
• Published: United States 2008
• Subjects: Animals; Cell Culture Techniques - methods; Cell Differentiation; Cell Survival; Cryopreservation; Dissection; Immunohistochemistry; Mice; Neurons - cytology; Rats; Stem Cells - cytology
• ISSN: 1064-3745
• DOI: 10.1007/978-1-59745-133-8_4

Recent work shows that major developmental and clinical processes such as central nervous system regeneration and carcinogenesis involve stem cells (SCs) in the brain. In spite of this importance, the requirements of these SCs and their differentiated offspring (neurons, astrocytes, and oligodendrocytes) for survival and proper function are little understood. In vivo, the SCs themselves interact with their environment. This "SC niche" may be complex because it likely includes cells of the vascular and immune systems. The ability to maintain (1) and differentiate (1 -4) central nervous system (CNS) SCs in tissue culture where they can be pharmacologically or genetically (5) manipulated provides a powerful starting point for understanding their behavior. We present detailed information on the methods that permit CNS SCs to differentiate into functional neurons in tissue culture. Important aspects of the culture systems include (1) homogeneity, so that the input and output of a manipulation is known to involve the SC itself; (2) growth in monolayer to visualize and study individual SCs and their offspring; and (3) the use of fully defined culture components to exclude unknown factors from the culture. These conditions support the differentiation of functional, electrically active neurons. These methods allow cell growth and differentiation from normal adult and diseased tissue derived from both animal models and clinical samples. Ultimate validation of such a system comes from accurate prediction of in vivo effects, and the methods we present for CNS SC culture have also successfully predicted regenerative responses in the injured adult nervous system.