Rapid recycling of glutamate transporters on the astroglial surface
Glutamate uptake by astroglial transporters confines excitatory transmission to the synaptic cleft. The efficiency of this mechanism depends on the transporter dynamics in the astrocyte membrane, which remains poorly understood. Here, we visualise the main glial glutamate transporter GLT1 by generating its pH-sensitive fluorescent analogue, GLT1-SEP. FRAP-based imaging shows that 70-75% of GLT1-SEP dwell on the surface of rat brain astroglia, recycling with a lifetime of ~22 s. Genetic deletion of the C-terminus accelerates GLT1-SEP membrane turnover while disrupting its surface pattern, as revealed by single-molecule localisation microscopy. Excitatory activity boosts surface mobility of GLT1-SEP, involving its C-terminus, metabotropic glutamate receptors, intracellular Ca2+ and calcineurin-phosphatase activity, but not the broad-range kinase activity. The results suggest that membrane turnover, rather than lateral diffusion, is the main 'redeployment' route for the immobile fraction (20-30%) of surface-expressed GLT1. This finding reveals an important mechanism helping to control extrasynaptic escape of glutamate.
All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for each corresponding Figure.
Article and author information
Wellcome Trust (212251_Z_18_Z)
- Dmitri A Rusakov
H2020 European Research Council (857562)
- Dmitri A Rusakov
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Animal experimentation: Animal expeirments were carried out in accordance with the national guideline and the European Communities Council Directive 0f November 1986, the European Directive 2010/63/EU on the Protection of Animals used for Scientific Purposes, and the United Kingdom Home Office (Scientific Procedures) Act of 1986, under UK Home Office Project licence PPL707524.
- Yukiko Goda, RIKEN, Japan
- Received: November 11, 2020
- Accepted: April 15, 2021
- Accepted Manuscript published: April 16, 2021 (version 1)
- Version of Record published: April 27, 2021 (version 2)
© 2021, Michaluk et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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