Abstract
Perisynaptic astrocytes express important glutamate transporters, especially excitatory amino acid transporter 2 (EAAT2, rodent analog GLT1) to regulate extracellular glutamate levels and modulate synaptic activation. In this study, we investigated an exciting new pathway, the exosome-mediated transfer of microRNA (in particular, miR-124a), in neuron-to-astrocyte signaling. Exosomes isolated from neuron-conditioned medium contain abundant microRNAs and small RNAs. These exosomes can be directly internalized into astrocytes and increase astrocyte miR-124a and GLT1 protein levels. Direct miR-124a transfection also significantly and selectively increases protein (but not mRNA) expression levels of GLT1 in cultured astrocytes. Consistent with our in vitro findings, intrastriatal injection of specific antisense against miR-124a into adult mice dramatically reduces GLT1 protein expression and glutamate uptake levels in striatum without reducing GLT1 mRNA levels. MiR-124a-mediated regulation of GLT1 expression appears to be indirect and is not mediated by its suppression of the putative GLT1 inhibitory ligand ephrinA3. Moreover, miR-124a is selectively reduced in the spinal cord tissue of end-stage SOD1 G93A mice, the mouse model of ALS. Subsequent exogenous delivery of miR-124a in vivo through stereotaxic injection significantly prevents further pathological loss of GLT1 proteins, as determined by GLT1 immunoreactivity in SOD1 G93A mice. Together, our study characterized a new neuron-to-astrocyte communication pathway and identified miRNAs that modulate GLT1 protein expression in astrocytes in vitro and in vivo.
Highlights
Pathways involved in neuron-dependent GLT1 regulation in astrocytes remain to be characterized
As miR-124a can be transferred from neurons to astrocytes through exosomes, we investigated whether miR-124a regulates GLT1 protein expression in astrocytes
We examined the effect of miR-124a-A/S on other glutamate transporters in the CNS, including the astroglial GLAST and neuronal EAAC1, neither of which has 3ЈUTR binding sites for miR-124a according to the TargetScan program
Summary
Pathways involved in neuron-dependent GLT1 regulation in astrocytes remain to be characterized. Differential translation efficiency among EAAT2/GLT1 mRNA transcripts has been found in an immortalized astroglial cell line following treatment with corticosterone or retinol [12] All these EAAT2/ GLT1 mRNA transcripts have a uniquely long 3Ј untranslated region (UTR) (9.7kb) [3, 13], implicating potential 3ЈUTR-dependent regulation, especially through microRNA-mediated translational inhibition or mRNA degradation. A recent study has demonstrated that genetic ablation of the neuronal EphA4 receptor results in a dramatic increase in GLT1 protein (but not mRNA) levels in the hippocampus [17], implicating a neuronal role in translational regulation of GLT1 expression. We investigated the transfer of neuronal exosomal miRNA (especially miR-124a) into astrocytes and miRNA-mediated translational regulation of GLT1 protein expression
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