Purification and culture of nerve growth factor receptor (p75)-expressing basal forebrain cholinergic neurons.
The activity of the basal forebrain cholinergic neurons (BFCNs) that innervate the cerebral cortex and hippocampus is essential for normal learning and memory. Here, we present a method to isolate and culture BFCNs from the embryonic murine septum that takes advantage of their restricted expression of the nerve growth factor receptor (p75) in conjunction with fluorescence-activated cell sorting. The septal region dissection, cell dissociation and staining process, and cell sorting parameters are described in detail. Sufficient cell yield and optimized cell culture conditions make this protocol suitable for multiple assays including immunocytochemistry, reverse transcriptase PCR, microarray profiling, acetylcholine measurements and electrophysiological assessment. The study of these neurons as a purified population will greatly advance our understanding of factors that influence their development and maintenance.
Schnitzler
Aletta C
AC
Department of Pathology and Laboratory Medicine, Boston University School of Medicine, Boston, Massachusetts 02118, USA.
Lopez-Coviella
Ignacio
I
Blusztajn
Jan Krzysztof
JK
D000818:Animals; D018929:Cell Culture Techniques; D002469:Cell Separation; D004210:Dissection; D005434:Flow Cytometry; D051379:Mice; D008815:Mice, Inbred Strains; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D020800:Receptor, Nerve Growth Factor / Q000378:metabolism*; D020665:Septum of Brain / Q000166:cytology* / Q000196:embryology / Q000378:metabolism
D020800:Receptor, Nerve Growth Factor