Cholesterol deficiency perturbs actin signaling and glutamate homeostasis in hippocampal astrocytes.

This study was undertaken to explore the possibility that cholesterol deficiency may perturb the physiological functions of astrocytes, thus rendering cells vulnerable to the cytotoxicity induced by glutamate (Glu). Cholesterol deprivation induces astrocyte stellation, which is accompanied by disruption of cortical actin, and phosphorylation of extracellular signal-regulated kinase (ERK) in an astrocyte-specific manner. Moreover, cholesterol reduction decreases the activity of glutamine synthetase (GS) while enhancing the capacity of Glu transporter. Using [(3)H]d-aspartate as a tracer, we found a marked efflux of [(3)H]d-aspartate from cholesterol-deficient astrocytes after Glu stimulation. Changes in the actin cytoskeleton, cell morphology, ERK phosphorylation and GS level gradually recovered in astrocytes after the withdrawal of cholesterol depletion. Moreover, withdrawal of cholesterol deprivation attenuated cell loss in cholesterol-deficient astrocytes during Glu exposure. Taken together, our data suggest that, upon Glu exposure, there would be an increase in intracellular Glu as a consequence of enhanced Glu uptake and reduced degradation of Glu by GS in cholesterol-deficient astrocytes. This in turn leads to a concentration gradient favoring Glu release, thereby causing the accumulation of cytotoxic levels of Glu extracellularly. It is thus concluded that the detrimental effect of cholesterol deprivation may, in part, arise from the impairment in Glu homeostasis.

Cholesterol deficiency increases the vulnerability of hippocampal glia in primary culture to glutamate-induced excitotoxicity.

Cholesterol, a molecule critical for cellular function, is found in particular high concentration in the brain and has been implicated to synaptic plasticity and neuronal regeneration. This study was undertaken to investigate the mechanism by which cholesterol shortage modulates glutamate (Glu)-induced excitotoxicity in hippocampal cell cultures. A combined treatment of lovastatin and beta-cyclodextrin reduced cellular content of cholesterol while having no significant effect on cell viability in neuron/glia mixed cultures. The experimental manipulation, nonetheless, exacerbated Glu-induced membrane damage and loss of mitochondrial activity in mixed cultures. Analysis of [3H]thymidine incorporation revealed cholesterol deficiency impaired cell proliferation in mixed cultures after Glu exposure, indicating considerable loss of glia. Indeed, it was found that cholesterol deprivation potentiated the release of lactate dehydrogenase (LDH) and the impairment in mitochondrial reduction of WST-1 reagent in astrocyte-enriched cultures subjected to Glu exposure. The detrimental effect of cholesterol shortage, nevertheless, was not observed in cultured neurons. Notably, the pretreatment of lovastatin and beta-cyclodextrin caused a decrease in the content of cellular LDH while having no effect on cell cycle profile and cellular activity of WST-1 reduction in astrocyte-enriched cultures. In contrast, removal of cholesterol had no effect on LDH content in neuron-enriched cultures. It is concluded that the differential vulnerability of cholesterol-depleted neural cells to excitotoxic damage may, in part, be ascribed to cholesterol shortage destabilizing the plasma membrane of astrocytes, thus rendering them less capable of withstanding Glu insult.