Receptor‐Mediated Phosphorylation of Astroglial Intermediate Filament Proteins in Cultured Astroglia

Abstract

Primary cultures of purified [rat] astroglia exhibit a variety of membrane receptors that regulate intracellular cAMP levels. The effect of such receptor angonists on protein phosphorylation were examined in intact astroglia. An analysis of 32P-labeled proteins derived from whole cell extracts and separated via 2-dimensional gel electrophoresis indicated that increasing cAMP levels in astroglia stimulated the phosphorylation of 2 distinct proteins that had apparent MW/isoelectric points (pI) of 51K/6.0 and 57K/5.7. Similar experiments with cultured meningeal cells indicated that only the 57K/5.7 protein was phosphorylated in response to elevated levels of cAMP. The 51K/6.0 protein was never observed in gels derived from meningeal cells. Immunoblot experiments indicated that the 51K/6.0 protein stained with antiserum to glial fibrillary acidic protein (GFAP) and the 57K/5.7 protein stained with antibodies to vimentin. Concentration-effect studies indicate that these proteins are maximally phosphorylated at concentrations of receptor agonists that only slightly elevate cAMP levels. All receptor agonists that increase cAMP levels appear similarly efficacious with respect to increasing the phosphorylation of the 2 proteins. Evidently, the membrane receptors present on astroglia function, in part, to regulate phosphorylation of the intermediate filament proteins GFAP and vimentin.