Abstract
Rett syndrome (RTT) is a neurodevelopmetal disorder associated with mutations in the methyl-CpG–binding protein 2 (MeCP2) gene. MeCP2-deficient mice recapitulate the neurological degeneration observed in RTT patients. Recent studies indicated a role of not only neurons but also glial cells in neuronal dysfunction in RTT. We cultured astrocytes from MeCP2-null mouse brain and examined astroglial gene expression, growth rate, cytotoxic effects, and glutamate (Glu) clearance. Semi-quantitative RT-PCR analysis revealed that expression of astroglial marker genes, including GFAP and S100β, was significantly higher in MeCP2-null astrocytes than in control astrocytes. Loss of MeCP2 did not affect astroglial cell morphology, growth, or cytotoxic effects, but did alter Glu clearance in astrocytes. When high extracellular Glu was added to the astrocyte cultures and incubated, a time-dependent decrease of extracellular Glu concentration occurred due to Glu clearance by astrocytes. Although the shapes of the profiles of Glu concentration versus time for each strain of astrocytes were grossly similar, Glu concentration in the medium of MeCP2-null astrocytes were lower than those of control astrocytes at 12 and 18 h. In addition, MeCP2 deficiency impaired downregulation of excitatory amino acid transporter 1 and 2 (EAAT1/2) transcripts, but not induction of glutamine synthetase (GS) transcripts, upon high Glu exposure. In contrast, GS protein was significantly higher in MeCP2-null astrocytes than in control astrocytes. These findings suggest that MeCP2 affects astroglial genes expression in cultured astrocytes, and that abnormal Glu clearance in MeCP2-deficient astrocytes may influence the onset and progression of RTT.
Highlights
Rett syndrome (RTT) is a neurodevelopmetal disorder that affects one in 15,000 female births, and represents a leading cause of mental retardation and autistic behavior in girls [1,2]
The relative expression levels of EAAT1 24 h after treatment were lower in the wild-type than in the methyl-CpG–binding protein 2 (MeCP2)-null culture, the difference was not statistically significant (Fig. 5D: 12 h; 1.10260.169 in control versus 1.09660.142 in MeCP2-null astrocytes, n = 6 each, p = 0.979, 24 h; 0.45660.123 in control versus 0.90160.172 in MeCP2-null astrocytes, n = 5 each, p = 0.068). These results suggest that MeCP2 deficiency affects the expression of glutamine synthetase (GS) and EAAT1 protein, and that accelerated Glu clearance may result from dysregulation of GS and EAAT1 protein in MeCP2-null astrocytes
We have demonstrated that MeCP2 regulates the expression of astroglial marker transcripts, including glial fibrillary acidic protein (GFAP) and S100b in cultured astrocytes
Summary
Rett syndrome (RTT) is a neurodevelopmetal disorder that affects one in 15,000 female births, and represents a leading cause of mental retardation and autistic behavior in girls [1,2]. Several studies using in vitro cell culture systems indicate that MeCP2 may play a role in processes of neuronal maturation including dendritic growth, synaptogenesis, and electrophysiological responses [1,7]. These data support the idea that MeCP2 deficiency in neurons is sufficient to cause an RTT-like phenotype. Very recent studies have indicated that re-expression of MeCP2 in astrocytes of MeCP2-deficient mice significantly improves locomotion, anxiety levels, breathing patterns, and average lifespan, suggesting that astrocyte dysfunction may be involved in the neuropathology and characteristic phenotypic regression of RTT [13]
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