Growth factor upregulation of a phosphoinositide-coupled metabotropic glutamate receptor in cortical astrocytes

Abstract

CNS function depends on a capacity for plasticity during development, following injury, and in response to changing environmental conditions. Functional alterations in signal transduction pathways and in neurotransmitter receptor expression are possible mechanisms for the expression of such plasticity. In the present report, we demonstrate that exposure of astrocytes to specific growth factors alters both the functional activity and the protein levels of a specific glutamate receptor. Exposure of astrocytes to basic fibroblast growth factor, epidermal growth factor, or transforming growth factor-alpha produced marked increases in the ability of metabotropic glutamate receptor (mGluR) agonists to stimulate phosphoinositide hydrolysis. Using Western immunoblotting, we demonstrate that an increase in the levels of one of the phosphoinositide-coupled mGluR subtypes, mGluR5, accompanies the increased ability of mGluR agonists to stimulate phosphoinositide hydrolysis. In contrast, another phosphoinositide-coupled subtype of this receptor family, mGluR1 alpha, was not present at detectable levels in these cultures. The enhanced stimulation of phosphoinositide hydrolysis showed little sensitivity to pertussis toxin, and appeared to be selective to mGluR agonists, as there was not a similar increase in the ability of norepinephrine or carbachol to stimulate phosphoinositide hydrolysis. These findings demonstrate that expression of mGluRs in astrocytes is plastic, and indicate a novel pathway through which specific growth factors may selectively modulate neurotransmitter action.

The Metabotropic Glutamate Receptor mGluR5 Induces Calcium Oscillations in Cultured Astrocytes via Protein Kinase C Phosphorylation

“…Astrocyte cultures were performed according to the procedures described by Miller et al (1995) with a minor modification. In brief, the neocortex of Sprague-Dawley rat pups (3-4 days old) was dissected, treated with dispase I (0.4 mg/ml), dissociated by trituration, and plated at a density of 5 X 106 cells per 75-cm 2 flask in Dulbecco's modified Eagle's medium (DMEM; adjusted to pH 7.4 with 25 mM HEPES and 14.3 mM NaHCO 3) supplemented with 15% fetal calf serum (FCS), 1 mM pyruvate, and 2 mM glutamine.…”

“…The probe used for mGluR5 mRNA was the l,824-bp BglII fragment of the rat mGluR5 eDNA clone, which corresponds to the transmembrane region common to two splice forms of mGluR5 mRNA (Abe et al, 1992;Minakami et al, 1993). For western blot analysis, cells were rinsed and scraped into a lysis buffer (Miller et al, 1995). After homogenization in 0.32 M sucrose, nuclei were removed by low-speed centrifugation, and a crude membrane fraction was prepared by centrifugation at 35,000 g for 1 h. Western blotting was performed by electrophoresis on a 7.5% sodium dodecyl sulfate polyacrylamide gel according to the procedure described previously (Shigemoto et al, 1993).…”

“…Several studies have indicated that glutamate elicits oscillatory [Ca 2~] 1responses in cultured astrocytes through [Ca 2~]mobilization (Cornell-Bell et al, 1990a;Glaum et al, 1990;de Barry et al, 1991;Holzwarth et al, 1994). It has also recently been reported that mGluR5 is preferentially expressed in cultured astrocytes and that this expression is up-regulated by specific growth factors such as basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), and transforming growth factor-a (TGF-a) (Miller et al, 1995). In this investigation, we examined whether mGluR5 is capable of inducing Ca2~I~oscillations in cultured astrocytes and, if so, whether PKC phosphorylation is crucial for mGluR5-mediated [Ca2~] 1oscillations.…”

Cyclothiazide selectively inhibits mGluR1 receptors interacting with a common allosteric site for non-competitive antagonists

“…In contrast to granule neurons, CTZ failed to inhibit PI hydrolysis stimulated by ACPD in cortical astrocytes (Fig. 7D), which express predominantly mGluR5 receptors (Miller et al, 1995).…”

Tissue specific differential response in metabolic activities of cerebrum, cerebellum, pons and medulla associated with thyroid dysfunction in sub-acute fluoride toxicity

“…Other than brain, consumption of fluoride through drinking water may cause adverse effect on other soft tissue such as thyroid, causing tremendous distress on its function [56]. Brain development at neonatal as well as growing stages is largely regulated by thyroid hormones [57,58], which also facilitate the maturation of granule neurons [59] and protect them from apoptosis [60]. Deficiency of thyroid hormones during the sensitive period of neurogenesis and neuronal migration can cause irremediable damage to various structures, leading to death of granule cells [61,62], and blunted dendritic arborisation of Purkinje cells [63,64].…”