Abstract
Dysregulation of the astroglial glutamate transporters GLAST and GLT-1 has been implicated in several neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS) where a loss of GLT-1 protein expression and activity is reported. Furthermore, the two principal C-terminal splice variants of GLT-1 (namely GLT-1a and GLT-1b) show altered expression ratio in animal models of this disease. Considering the putative link between inflammation and excitotoxicity, we have here characterized the influence of TNF-α on glutamate transporters in cerebral cortical astrocyte cultures from wild-type rats and from a rat model of ALS (hSOD1G93A). Contrasting with the down-regulation of GLAST, a 72 h treatment with TNF-α substantially increased the expression of GLT-1a and GLT-1b in both astrocyte cultures. However, as the basal level of GLT-1a appeared considerably lower in hSOD1G93A astrocytes, its up-regulation by TNF-α was insufficient to recapitulate the expression observed in wild-type astrocytes. Also the glutamate uptake activity after TNF-α treatment was lower for hSOD1G93A astrocytes as compared to wild-type astrocytes. In the presence of the protein synthesis inhibitor cycloheximide, TNF-α did not influence GLT-1 isoform expression, suggesting an active role of dynamically regulated protein partners in the adaptation of astrocytes to the inflammatory environment. Confirming the influence of inflammation on the control of glutamate transmission by astrocytes, these results shed light on the regulation of glutamate transporter isoforms in neurodegenerative disorders.
Highlights
Disturbances in glutamate homeostasis, which lead to toxic accumulation of this excitatory neurotransmitter in the synaptic cleft, are observed in several neuropathologies
While cortical astrocyte cultures derived from newborn rodents are widely used to study specific astrocytic alteration associated with diverse insults, the expression of the human mutated form of SOD1 has never been characterized in cultures derived from hSOD1G93A rat pups
The rSOD1 protein was detected in astrocytes from both wild-type and transgenic rats, even though the signal corresponding to this endogenous protein is considerably lower as compared to the human protein in the hSOD1G93A samples
Summary
Disturbances in glutamate homeostasis, which lead to toxic accumulation of this excitatory neurotransmitter in the synaptic cleft, are observed in several neuropathologies. Among the five subtypes of glutamate transporters identified in cells of the nervous parenchyma, the astroglial glutamate transporters GLAST (the rodent homologue of human EAAT1) and GLT-1 (EAAT2) [2] are best characterized as they likely ensure most of the glutamate uptake in physiological conditions. Dysfunction of these two glutamate transporters is clearly implicated in neurodegenerative insults, in particular amyotrophic lateral sclerosis (ALS) [3,4], a fatal disease characterized by the progressive and specific loss of motor neurons that has been correlated with deficient GLT-1 expression and activity [5,6]. Noteworthy that so far, the majority of biochemical studies failed to distinguish between these two isoforms or essentially focused on GLT-1a because the distinctive roles of each of these isoforms and their physiological relevance remain poorly understood
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