Characterization of functional nerve growth factor‐receptors in a CNS glial cell line: Monoclonal antibody 217c recognizes the nerve growth factor‐receptor on C6 glioma cells

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Abstract
The biological effects of nerve growth factor (NGF) have been shown to be mediated by the high‐affinity form of the nerve growth factor receptor (NGF‐R) in sympathetic and sensory neurons, and in PC12 cells. We report here that the central nervous system C6 rat glioma cell line likewise expresses functional high‐affinity NGF‐Rs. The expression of NGF‐R mRNA in C6 cells can be up‐regulated by cycloheximide and its own ligand, NGF; and it can be rapidly down‐regulated by epidermal growth factor (EGF). Furthermore, C6 cells display NGF responsiveness by expressing c‐fos mRNA within 30 minutes of treatment with NGF; and after 4—5 days of NGF exposure, C6 cells cease dividing as measured by [3H]‐thymidine uptake, change shape, and reveal neurite‐like ‐processes. Scatchard analysis of [125 I]‐labelled bound to solubilized C6 cells confirms the presence of both high‐ and low‐affinity receptor protein. Cross‐linking radiolabeled NGF to its receptor in the presence or absence of excess unlabeled NGF, followed by immunoprecipitation with monoclonal antibody (mAb) 192‐IgG (a known anti‐NGF‐R antibody) and SDS‐PAGE reveals a 100 kD band corresponding to the NGF/NGF‐R complex. An identical band is observed when the immunoprecipitation is carried out with mAb 217c, suggesting that the 217c epitope is related to NGF‐R. The 217c antibody was generated against C6 cells and shown to be a cell surface antibody (Peng et al., Science 215:1102–4, 1982); several investigators have used it subsequently as an immunocytochemical marker for Schwann cells. The significance of NGF‐Rs in a CNS glial cell line is unclear, but association of NGF with the control of proliferation and/or differentiation of primitive glial cells is suggested.