Insulin and IGF-I stimulate phosphorylation of their respective receptors in intact neuronal and glial cells in primary culture.
Previous studies have shown that insulin and IGF-I bind to their respective receptors and stimulate autophosphorylation of the receptor beta subunits in detergent extracts of neuronal and glial cells. In the present study, intact neuronal and glial cells in primary culture have been utilized to characterize insulin- and IGF-I-stimulated phosphorylation of their receptors. Following [32P]orthophosphate labelling and stimulation by insulin or IGF-I, the cells were solubilized and the phosphorylated receptors were partially purified on wheat germ agglutinin--agarose columns, and immunoprecipitated using anti-phosphotyrosine or anti-insulin receptor antibodies. Insulin stimulated the phosphorylation of its receptor beta subunit (95 kD phosphoprotein) in a dose-dependent manner, within at least 20 seconds in both neuronal and glial cells. Additionally, a 102-kD phosphoprotein was observed in insulin-stimulated neuronal cells. Maximal stimulation of receptor phosphorylation occurred at 1 minute for the glial cells, and 10 minutes for the neuronal cells. IGF-I stimulated the phosphorylation of two phosphoproteins in intact neuronal and glial cells; a 95-kD protein and a 102-kD protein, in a dose-dependent manner. These observations demonstrate that both insulin and IGF-I stimulate the phosphorylation of the beta subunits of their respective receptors in brain cells in a similar fashion to their effects on receptors from nonneural tissues.
Shemer
J
J
Diabetes Branch, National Institute of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
Adamo
M
M
Raizada
M K
MK
Heffez
D
D
Zick
Y
Y
LeRoith
D
D
D000818:Animals; D000831:Animals, Newborn; D001921:Brain / Q000378:metabolism; D002478:Cells, Cultured; D007328:Insulin / Q000494:pharmacology*; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D009457:Neuroglia / Q000187:drug effects / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D011972:Receptor, Insulin / Q000187:drug effects / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000187:drug effects / Q000378:metabolism*; D017451:Receptors, Somatomedin; D011994:Recombinant Proteins / Q000494:pharmacology; D013002:Somatomedins / Q000494:pharmacology*
D007328:Insulin; D046911:Macromolecular Substances; D011956:Receptors, Cell Surface; D017451:Receptors, Somatomedin; D011994:Recombinant Proteins; D013002:Somatomedins; D007334:Insulin-Like Growth Factor I; D011972:Receptor, Insulin