PKC-Mediated Modulation of Astrocyte SNAT3 Glutamine Transporter Function at Synapses in Situ

Date

2018

Authors

Dong, Wuxing
Todd, Alison
Broer, Angelika
Hulme, Sarah
Broer, Stefan
Billups, Brian

Journal Title

Journal ISSN

Volume Title

Publisher

MDPI Publishing

Abstract

Astrocytes are glial cells that have an intimate physical and functional association with synapses in the brain. One of their main roles is to recycle the neurotransmitters glutamate and gamma-aminobutyric acid (GABA), as a component of the glutamate/GABA-glutamine cycle. They perform this function by sequestering neurotransmitters and releasing glutamine via the neutral amino acid transporter SNAT3. In this way, astrocytes regulate the availability of neurotransmitters and subsequently influence synaptic function. Since many plasma membrane transporters are regulated by protein kinase C (PKC), the aim of this study was to understand how PKC influences SNAT3 glutamine transport in astrocytes located immediately adjacent to synapses. We studied SNAT3 transport by whole-cell patch-clamping and fluorescence pH imaging of single astrocytes in acutely isolated brainstem slices, adjacent to the calyx of the Held synapse. Activation of SNAT3-mediated glutamine transport in these astrocytes was reduced to 77 ± 6% when PKC was activated with phorbol 12-myristate 13-acetate (PMA). This effect was very rapid (within ~20 min) and eliminated by application of bisindolylmaleimide I (Bis I) or 7-hydroxystaurosporine (UCN-01), suggesting that activation of conventional isoforms of PKC reduces SNAT3 function. In addition, cell surface biotinylation experiments in these brain slices show that the amount of SNAT3 in the plasma membrane is reduced by a comparable amount (to 68 ± 5%) upon activation of PKC. This indicates a role for PKC in dynamically controlling the trafficking of SNAT3 transporters in astrocytes in situ. These data demonstrate that PKC rapidly regulates the astrocytic glutamine release mechanism, which would influence the glutamine availability for adjacent synapses and control levels of neurotransmission.

Description

Keywords

Slc38a3, system N, protein kinase C, phorbol ester, protein trafficking, phosphorylation, biotinylation, glia, calyx of Held

Citation

Source

International Journal of Molecular Sciences

Type

Journal article

Book Title

Entity type

Access Statement

Open Access

License Rights

Creative Commons Attribution (CC BY) license

DOI

10.3390/ijms19040924

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