Characterization of epidermal growth factor receptors in astrocytic glial and neuronal cells in primary culture.
Studies characterized the structure and function of epidermal growth factor (EGF) receptors in astrocytic glial cells and neuronal cells in primary culture from neonatal rat brain. [125I]EGF binding to membranes prepared from glial and neuronal cultures was specific and dependent on protein concentration; however, glial preparations bound 5-fold more [125I]EGF per mg protein. Unlabeled EGF competed for binding to both glial and neuronal membranes with an IC50 of 5 nM, whereas insulin, insulin-like growth factor I, and nerve growth factor failed to compete. Scatchard plot analysis of binding data for glial cells yielded a curvilinear plot with dissociation constants of 7.12 nM for high affinity and 6.2 microM for low affinity sites. The higher level of binding in glial compared to neuronal membranes reflected a greater number of binding sites rather than differences in receptor affinity. In glial membranes, [125I]EGF covalently cross-linked to one major protein with a mol wt of 170,000, and EGF stimulated the phosphorylation of a 170,000 protein which was half-maximal at 20 nM. In contrast, neither covalent cross-linking nor receptor autophosphorylation could be detected in neuronal membranes. Culture of glial cells in the presence of EGF stimulated [35S]methionine incorporation into both cellular and secreted proteins, whereas no effect of EGF was observed in neuronal cultures. The addition of EGF to glial cultures produced a dose-dependent stimulation of [3H]thymidine incorporation as well as the multiplication of cells over a 6-day period. These observations show that functional EGF receptors in the neonatal brain are predominantly localized in glial cells.
Department of Pharmacology and Experimental Therapeutics, University of Florida, Gainesville 32610.
S L
SL
Wang
K T
KT
Shiverick
S
S
Ogilvie
W A
WA
Dunn
M K
MK
Raizada
D000818:Animals; D000831:Animals, Newborn; D001253:Astrocytes / Q000378:metabolism*; D001667:Binding, Competitive; D001921:Brain / Q000166:cytology; D002455:Cell Division; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D003432:Cross-Linking Reagents; D004815:Epidermal Growth Factor / Q000378:metabolism / Q000494:pharmacology; D066246:ErbB Receptors / Q000378:metabolism*; D008970:Molecular Weight; D009419:Nerve Tissue Proteins / Q000096:biosynthesis; D009474:Neurons / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats
D003432:Cross-Linking Reagents; D009419:Nerve Tissue Proteins; D004815:Epidermal Growth Factor; D066246:ErbB Receptors