Elsevier

Neuropharmacology

Volume 62, Issue 2, February 2012, Pages 797-806
Neuropharmacology

NMDA GluN2A and GluN2B receptors play separate roles in the induction of LTP and LTD in the amygdala and in the acquisition and extinction of conditioned fear

https://doi.org/10.1016/j.neuropharm.2011.09.001Get rights and content

Abstract

Synaptic plasticity mediated by NMDA glutamate receptors is thought to be a primary mechanism underlying the formation of new memories. Activation of GluN2A NMDA receptor subunits may induce long-term potentiation (LTP), whereas low-frequency stimulation of GluN2B receptors induces long-term depression (LTD). In the present study, we show that blockade of GluN2A, but not GluN2B receptors with NVP-AAM077 and Ro25-6981 respectively, prevented LTP of auditory thalamic inputs to the lateral amygdala. Conversely, LTD induction in this pathway was prevented by blockade of GluN2B, but not GluN2A receptors. As this pathway plays a critical role in the acquisition, retrieval and extinction of a learned auditory-cue fear association, we next examined the effects of blockade of GluN2A and GluN2B receptors on the development and retention of a conditioned fear response. Administration of NVP-AAM077, but not Ro25-6981, prior to conditioning disrupted the expression of conditioned fear 24 h later. Conversely, Ro25-6981 but not NVP-AAM077 impaired extinction of the conditioned fear response. These data expand on previous work showing that LTP/D in the thalamic-lateral amygdala pathway is dependent on NMDA receptors, by demonstrating selective roles for GluN2A and GluN2B NMDA receptor subunits in LTP and LTD respectively. Furthermore, GluN2A receptor activation and associated LTP may be involved specifically in the initial formation and/or stabilization of a learned fear response, whereas GluN2B receptor activation and associated LTD may facilitate the suppression of Pavlovian fear responses during extinction.

This article is part of a Special Issue entitled ‘Post-Traumatic Stress Disorder’.

Highlights

► NMDA GluN2A/B receptor subtypes dissociably facilitate LTP/LTD in the amygdala. ► Blockade of GluN2A receptors selectively weaken acquisition of fear memories. ► Disruption of GluN2B receptors selectively retards extinction of fear memories.

Introduction

Changes in the strength of synaptic transmission provide a fundamental mechanism through which learning and memory occurs in the mammalian brain. Long-term potentiation (LTP) and depression (LTD) of synaptic activity are currently the best characterized forms of synaptic plasticity and are suggested as physiological substrates of learning and memory (Bliss and Collingridge, 1993, Bear and Malenka, 1994, Rioult-Pedotti et al., 1998, Whitlock et al., 2006).

Fear conditioning is a simple form of associative learning that entails an association between a previously neutral conditioned stimulus (CS) with the onset/occurrence of an aversive unconditioned stimulus (US). After repeated pairings, presentation of the CS alone elicits behavioral and physiological responses consistent with fear or anxiety (Fendt and Fanselow, 1999, Gewirtz and Davis, 2000, LeDoux, 2000, Nijsen et al., 1998). LTP-like changes have been observed following the formation of a CS/US association (McKernan and Shinnick-Gallagher, 1997, Rogan and LeDoux, 1995, Rogan et al., 1997) and treatments that block the induction of LTP in vitro prevent the formation of a Pavlovian fear response in rats (Bauer et al., 2002). Although the mechanisms through which Pavlovian fear is extinguished are still debated, recent evidence suggests that a reversal of learning-induced potentiation and/or LTD may play an important role (Dalton et al., 2008, Kim et al., 2007). In a seminal study, Quirk et al. (1995) demonstrated that neurons of the lateral amygdala (LA) showed a significant increase in firing in response to a tone CS following pairing with a footshock US. This LTP-like response was attenuated following extinction training such that firing of these neurons resembled pre-conditioning levels once extinction was achieved. Subsequently, Lin et al. (2003) demonstrated that depotentiation of LA neurons with low-frequency stimulation administered after fear conditioning blocked the expression of conditioned fear, in a manner resembling extinction.

Recent work from our laboratory has examined the role of LTD on fear learning and extinction. Administration of an interference peptide (Tat-GluR23Y), that blocks AMPA receptor endocytosis and LTD in vivo (Brebner et al., 2005, Fox et al., 2007, Wong et al., 2007), selectively blocked the extinction of a learned fear response (Dalton et al., 2008). This effect is particularly interesting given that the same peptide had no effect on the acquisition of a learned fear response in these animals. These results were replicated by Kim et al. (2007) using intra-amygdala infusions of the same peptide. These data suggest that LTD may be a primary mechanism through which inappropriate responses are suppressed while mechanisms responsible for the acquisition of new learning remain intact. This hypothesis is supported by a recent report that the selective GluN2B N-methyl-d-aspartate (NMDA) receptor subunit antagonist Ro25-6981 not only blocked the induction of LTD but also increased perseveration during reversal learning in a water maze (Duffy et al., 2008), increased premature responding in a 5-choice serial reaction time task (Higgins et al., 2003), increased perseveration while shifting strategies in a set-shifting paradigm (Dalton et al., 2011), and induced a similarly selective impairment in extinction learning to Tat-GluR23Y (Dalton et al., 2008). These data are consistent with a role for LTD in the inhibition of a previously learned response.

Collectively, these findings support the conjecture that LTP-like processes play a critical role in the acquisition of a new response (such as learned fear) whereas LTD-like processes may contribute to the suppression of a previously acquired but no longer appropriate response (as occurs during extinction). Until recently, appropriate pharmacological means with which to selectively target each of these processes were lacking. Recent electrophysiological data suggest a functional dissociation between the activation of a given NMDA receptor subunit and the induction of LTP as distinct from LTD. Under certain experimental conditions, GluN2A subunit activation results in the induction of an LTP-like response, whereas activation of receptors containing the GluN2B subunit mediates the generation of LTD (Izumi et al., 2006, Liu et al., 2004, Woo et al., 2005, Yang et al., 2005; but see Hendricson et al., 2002, Morishita et al., 2007). Thus, in the present study, we investigated the potential roles of GluN2A or GluN2B receptors in 1) the induction of LTP or LTD in the LA, a nucleus critical for fear memories and 2) the acquisition and extinction of associative fear memories.

Section snippets

Experiment 1: relative contribution of GluN2A and GluN2B receptors to induction of LTP and LTD in the LA

Male Sprague Dawley rats (17–24 days old; Charles River Laboratories) were placed under deep anesthesia and decapitated. The brain was rapidly removed into ice-cold slicing solution containing (in mM): 87 NaCl, 2.5 KCl, 7 MgCl2, 0.5 CaCl2, 1.25 NaH2PO4, 26 NaHCO3, 25 glucose and 75 sucrose that infused continuously with carbogen (95% O2 and 5% CO2) to adjust pH to 7.4. Coronal slices of 400 μm thickness containing the amygdala were produced using a vibrating blade microtome and recovered in an

Experiment 1: differential roles of GluN2A- and GluN2B-containing NMDA receptors in the LTP and LTD at thalamic input synapses in the LA

We employed standard pairing protocols to induce LTP and LTD in pyramidal neurons of the LA. Our initial experiments on synaptic plasticity within this region revealed that both LTP and LTD were blocked by the non-specific NMDA-receptor antagonist, APV (Yu et al., 2008). Subsequent experiments that used identical procedures and are described here were designed to ascertain the relative contribution of different NMDA receptor subtypes to these forms of plasticity.

As shown in Fig. 1, stimulation

Discussion

The major findings of this study concern important differences between the effects of GluN2A and GluN2B receptor blockade on both the induction of LTP or LTD and acquisition and extinction of conditioned fear. It has been well established that synaptic plasticity in this region is critically dependant on NMDA receptor activity. Here we show for the first time that NVP, a GluN2A receptor-preferring antagonist, disrupted both LTP in amygdala slices and also compromised the acquisition of a

Acknowledgements

This work was supported by grants from the Natural Sciences and Engineering Research Council of Canada to AGP and SBF and the Canadian Institutes of Health Research to AGP and YTW. SBF is a Michael Smith Foundation for Health Research Senior Scholar. We thank Dr. Y. P. Auberson (Novartis Pharma AG, Basel, Switzerland) for the generous gift of NVP-AAM077.

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