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

Abstract

Primary cultures of purified astroglia have been shown to exhibit a variety of membrane receptors that regulate intracellular cyclic AMP levels. The experiments described in this paper were completed to examine the effect of such receptor agonists on protein phosphorylation in intact astroglia. An analysis of 32P-labelled proteins derived from whole cell extracts and separated via two-dimensional gel electrophoresis indicated that increasing cyclic AMP levels in astroglia stimulated the phosphorylation of two distinct proteins that had apparent molecular weights/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 cyclic AMP. 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 cyclic AMP levels. All receptor agonists that have been shown to increase cyclic AMP levels appear similarly efficacious with respect to increasing the phosphorylation of the two proteins. These experiments suggest that the membrane receptors present on astroglia function, in part, to regulate phosphorylation of the intermediate filament proteins GFAP and vimentin.

Neuronal modulation of schwann cell glial fibrillary acidic protein (GFAP)

“…and Riuna, 1984;McCarthy et al, 1985;Fields and McMenamin, 1985) or other modes of posttranslational processing of these peptides in CNS and PNS. An alternate possibility is that CNS and PNS GFAP peptides are translated from GFAP mRNAs that, although approximately the same size, have been alternatively spliced.…”

Human immunodeficiency virus coat protein gp120 inhibits the beta-adrenergic regulation of astroglial and microglial functions.

“…receptors, among which 3-adrenergic receptors are among the most studied and the most effective (14,15). Table 1 shows the massive (up to 90-fold), dose-dependent elevation in cAMP levels induced by a 10-min exposure to the -3-adrenergic agonist isoproterenol (1 nM-1 1.M) or to the adenylate cyclase activator forskolin (30 and 100 1.M) in cultured rat astrocytes.…”

“…In astrocytes, the activation of 3-adrenergic receptors was shown to stimulate the phosphorylation of the two major cytoskeletal proteins vimentin and GFAP (14), to induce the secretion ofnerve growth factor (16) and of the neuroactive amino acid taurine (15), and to down-regulate the expression of major histocompatibility complex class II antigens (17). In activated microglia, (-adrenergic receptors were reported to modulate the expression and secretion of inflammatory cytokines, and in particular to down-regulate the production of TNF-a (19), probably through a mechanism involving cAMP (20).…”

Regulation of Neuregulin Expression in the Injured Rat Brain and Cultured Astrocytes

“…Thus, candidate regulators of neuregulin expression were chosen on the basis of their previously published effects on the expression of the various neurotrophic factors in astrocyte cultures (Condorelli et al, 1994;Rudge et al, 1994). Because a transient increase in the intracellular cAMP level induces astrocyte differentiation, including morphological changes, increased GFAP expression, and increased phosphorylation of intermediate filament proteins (Sensenbrenner et al, 1980;McCarthy et al, 1985), we selected cAMP-elevating reagents to investigate the regulation of neuregulin expression. Figure 6 B shows that forskolin and cAMP analogs enhanced the expression level of the 52 kDa neuregulin on astrocyte cell surfaces after a 3 hr incubation.…”