Elsevier

Neuropharmacology

Volume 41, Issue 6, November 2001, Pages 664-671
Neuropharmacology

Metabotropic glutamate receptor activation causes a rapid redistribution of AMPA receptors

https://doi.org/10.1016/S0028-3908(01)00134-4Get rights and content

Abstract

Electrophysiology, immunostaining and time lapse imaging techniques were employed to study the mechanism of long-term depression (LTD) induced by DHPG, a specific group I metabotropic glutamate receptor (mGluR) agonist. Experiments were performed in primary hippocampal culture or in the CA1 area of acute rat hippocampal slices. In agreement with previous results by others, we show that DHPG (200 μM, 10 min) can induce LTD (DHPG-LTD) in acute slices, in the presence or absence of synaptic inhibition. In addition, in voltage clamp whole cell experiments we find that accompanying the reduction in the evoked excitatory postsynaptic current (EPSC), miniature EPSC amplitude and frequency are reduced. Similar results were obtained in cultured neurons. Immunostaining and time lapse imaging showed a long-lasting loss of AMPA receptors from the membrane surface of cultured neurons after DHPG treatment, which appears to occur in only a subset of the puncta. Further electrophysiological recordings on slices showed that blocking postsynaptic endocytosis by introducing a blocking peptide named D15 in recording pipettes abolished the DHPG-LTD. In conclusion, these data suggest that LTD induced by mGluR activation is due to a rapid removal of AMPA receptors from the postsynaptic membrane.

Introduction

It is well established that at many excitatory synapses repetitive stimulation leads to long lasting changes in synaptic strength. Brief high frequency stimulation results in long term potentiation (LTP), whereas prolonged low frequency stimulation (LFS) results in long term depression (LTD). Under most conditions both LTP and LTD are dependent on the activation of NMDA receptors (Bear and Malenka, 1994, Bear and Abraham, 1996). However, there are a number of reports suggesting that a metabotropic glutamate receptor dependent form of LTD (mGluR-LTD) also exists in the CA1 region of the hippocampus (Stanton et al., 1991, Bolshakov and Siegelbaum, 1994, Yang et al., 1994, Oliet et al., 1997) and may coexist at excitatory synapses with the NMDA receptor dependent form of LTD (NMDAR-LTD) (Oliet et al., 1997). Recent evidence suggests that selective removal of AMPA receptors from the postsynaptic membrane (Carroll et al., 1999, Lüscher et al., 1999, Lüthi et al., 1999, Man et al., 2000, Daw et al., 2000) may contribute to NMDAR-LTD. However, little is known about the underlying mechanisms of mGluR-LTD.

The mGluR-LTD induced by LFS requires activation of group I mGluRs and is associated with a large decrease of miniature EPSC (mEPSC) frequency (Oliet et al., 1997). This could be due either to a presynaptic change, e.g. reduction in transmitter release, or a postsynaptic change, e.g. removal of AMPA receptors from postsynaptic membrane surface. Group I mGluRs, and in particular mGluR5s, are located postsynaptically on CA1 pyramidal cells (Lujan et al., 1996, Lujan et al., 1997) and application of a selective group I mGluR agonist, DHPG, can induce a long lasting depression of synaptic transmission (DHPG-LTD) (Palmer et al., 1997, Fitzjohn et al., 1999, Schnabel et al., 1999, Huber et al., 2000), which requires postsynaptic protein synthesis (Huber et al., 2000). Although DHPG-LTD shares a number of features with mGluR-LTD activated by LFS, it is not known whether the expression of either form of LTD is a pre or postsynaptic phenomenon.

To address this issue, we carried out a series of experiments both in slice and culture preparations, using electrophysiology, immunostaining and time lapse imaging techniques. In agreement with previous results, we show that DHPG can induce LTD in acute slices, in the presence or absence of synaptic inhibition. In addition, we find that accompanying the reduction in the evoked EPSC, mEPSC amplitude and frequency are reduced. Similar results were obtained in cultured neurons. Immunostaining and time lapse imaging showed a long-lasting loss of AMPA receptors from the membrane surface of cultured neurons after DHPG treatment, which appears to occur in only a subset of the synapses. Further electrophysiological recordings on slices showed blocking endocytosis by introducing a blocking peptide into the recording pipette abolished the DHPG-LTD. In all, our data suggest that the DHPG-LTD is due to removal of AMPA receptors, resembling NMDAR-LTD or the effects of ionotropic GluR agonists on glutamate receptors (e.g. glutamate, NMDA or AMPA).

Section snippets

Slice preparations

Slices were prepared from young (10–21 day old) Sprague-Dawley rats. In brief, rats were anesthetized with halothane and decapitated, and their brains were removed. The hippocampi were dissected out and transverse slices (300–400 μm) were prepared by a vibrotome Leica VT1000S (Leica Microsystems Nussloch GmbH, Germany), cut at 4°C artificial cerebrospinal fluid (ACSF) containing (in mM) NaCl 119, KCl 2.5, CaCl2 2.5, MgCl2 1.3, NaHCO3 26.2, NaH2PO4 1 and glucose 11(saturated with 95% O2 and 5% CO

Group I mGLuR agonist DHPG causes a long lasting depression of field EPSPs (DHPG-LTD) in hippocampal slices

In the first set of experiments, field EPSPs were recorded with inhibition intact. The slope of field EPSPs was measured and after 15 min of stable baseline, DHPG (200 μM) was applied for 10 min. This treatment caused an initial large reduction of field EPSP followed by a slow recovery, but the EPSPs remained at a depressed level more than 30 min after washout of the drug. The average value for the 4 min period measured at 21–25 min after DHPG washout was 79.8±3.3% (n=12, p<0.01, Fig. 1A). To

Discussion

The present electrophysiological results, in agreement with a number of previous reports (Palmer et al., 1997, Schnabel et al., 1999, Fitzjohn et al., 1999, Huber et al., 2000), show that a brief activation of group I mGluRs by the selective group I mGluR agonist DHPG induces a long lasting depression of synaptic transmission in the hippocampal CA1 area. Initially there is a large rapid depression in synaptic transmission during the application of DHPG and this is followed by a long lasting but

Acknowledgements

R.A.N. is supported by grants from the NIH and Bristol-Myers Squibb Co. and is a member of the Keck Center for Integrative Neuroscience and the Silvio Conte Center for Neuroscience Research.

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1

Present address: Department of Medical Biophysics, Institute of Physiology and Pharmacology, Gothenburg University, Medicinaregatan 1D, Box 433, Gothenburg, SE 405 30, Sweden.

2

Present address: Department of Molecular and Cell Biology, University of California at Berkeley, Berkeley, CA 94720, USA.

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