A novel purification method for CNS projection neurons leads to the identification of brain vascular cells as a source of trophic support for corticospinal motor neurons.
One of the difficulties in studying cellular interactions in the CNS is the lack of effective methods to purify specific neuronal populations of interest. We report the development of a novel purification scheme, cholera toxin beta (CTB) immunopanning, in which a particular CNS neuron population is selectively labeled via retrograde axonal transport of the cell-surface epitope CTB, and then purified via immobilization with anti-CTB antibody. We have demonstrated the usefulness and versatility of this method by purifying both retinal ganglion cells and corticospinal motor neurons (CSMNs). Genomic expression analyses of purified CSMNs revealed that they express significant levels of many receptors for growth factors produced by brain endothelial cells; three of these factors, CXCL12, pleiotrophin, and IGF2 significantly enhanced purified CSMN survival, similar to previously characterized CSMN trophic factors BDNF and IGF1. In addition, endothelial cell conditioned medium significantly promoted CSMN neurite outgrowth. These findings demonstrate a useful method for the purification of several different types of CNS projection neurons, which in principle should work in many mammalian species, and provide evidence that endothelial-derived factors may represent an overlooked source of trophic support for neurons in the brain.
Dugas
Jason C
JC
Department of Neurobiology, Stanford University School of Medicine, Stanford, California 94305-5125, USA.jcdugas@alum.mit.edu
Mandemakers
Wim
W
Rogers
Madolyn
M
Ibrahim
Adiljan
A
Daneman
Richard
R
Barres
Ben A
BA
D000818:Animals; D001370:Axonal Transport / Q000502:physiology; D001921:Brain / Q000098:blood supply / Q000166:cytology* / Q000502:physiology; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D002772:Cholera Toxin / Q000502:physiology; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D009046:Motor Neurons / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D009414:Nerve Growth Factors / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D009434:Neural Pathways / Q000166:cytology / Q000502:physiology; D009474:Neurons / Q000166:cytology / Q000378:metabolism / Q000502:physiology; D011712:Pyramidal Tracts / Q000098:blood supply / Q000166:cytology* / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley
D009414:Nerve Growth Factors; D002772:Cholera Toxin