Differential trafficking of adenosine receptors in hippocampal neurons monitored using GFP- and super-ecliptic pHluorin-tagged receptors

A. E. Baines, S. A. L. Correa, A.J. Irving, B. G. Frenguelli

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    Abstract

    Adenosine receptors (ARs) modulate many cellular and systems-level processes in the mammalian CNS. However, little is known about the trafficking of ARs in neurons, despite their importance in controlling seizure activity and in neuroprotection in cerebral ischaemia. To address this we examined the agonist-dependent internalisation of C-terminal GFP-tagged A(1)Rs, A(2A)Rs and A(3)Rs in primary hippocampal neurons. Furthermore, we developed a novel super-ecliptic pHluorin (SEP)-tagged AIR which, via the N-terminal SEP tag, reports the cell-surface expression and trafficking of A(1)Rs in real-time. We demonstrate the differential trafficking of ARs in neurons: A(3)Rs internalise more rapidly than A(1)Rs, with little evidence of appreciable A(2A)R trafficking over the time-course of the experiments. Furthermore, the novel SEP-A(1)R construct revealed the time-course of internalisation and recovery of cell-surface expression to occur within minutes of agonist exposure and removal, respectively. These observations highlight the labile nature of A(1)R and A(3)Rs when expressed at the neuronal plasma membrane. Given the high levels of adenosine in the brain during ischaemia and seizures, internalisation of the inhibitory A(1)R may result in hyperexcitability, increased brain damage and the development of chronic epileptic states. (C) 2011 Elsevier Ltd. All rights reserved.

    Original languageEnglish
    Pages (from-to)1-11
    Number of pages11
    JournalNeuropharmacology
    Volume61
    Issue number1-2
    DOIs
    Publication statusPublished - 2011

    Keywords

    • Adenosine receptor
    • Internalisation
    • Trafficking
    • Epilepsy
    • Seizure
    • Ischaemia
    • AGONIST-DEPENDENT PHOSPHORYLATION
    • TEMPORAL-LOBE EPILEPSY
    • POSTISCHEMIC RAT-BRAIN
    • A(2A) RECEPTOR
    • A(3) RECEPTORS
    • SYNAPTIC-TRANSMISSION
    • NERVOUS-SYSTEM
    • A(1) RECEPTORS
    • MULTIPLE MECHANISMS
    • CALCIUM-CHANNELS

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