Amygdalar circuits required for either consolidation or extinction of taste aversion memory are not required for reconsolidation

Recent reports have revitalized the debate on whether, for each item in memory, consolidation occurs just once, or whether, upon their activation in retrieval, items in memory undergo reconsolidation. Further, it has been recently reported that following retrieval in the absence of reinforcer, the activated memory can either reconsolidate or extinguish, depending on the training history. This raises the question whether consolidation, extinction and reconsolidation share neuronal mechanisms, and moreover, whether reconsolidation recapitulates consolidation. In conditioned taste aversion (CTA), consolidation depends on protein synthesis in the central nucleus of the amygdala, whereas extinction depends on protein synthesis in the basolateral nuclei of the amygdala. Here we show that inhibition of protein synthesis in either of these nuclei has no effect on CTA memory under conditions that initiate reconsolidation. This implies that reconsolidation does not recapitulate consolidation, and that consolidation, reconsolidation and extinction are different processes.     

Critical role of insular cortex in taste but not odour aversion memory

Conditioned odour aversion (COA) and conditioned taste aversion (CTA) result from the association of a novel odour or a novel taste with delayed visceral illness. The insular cortex (IC) is crucial for CTA memory, and the present experiments sought to determine whether the IC is required for the formation and the retrieval of COA memory as it is for CTA. We first demonstrated that ingested odour is as effective as taste for single-trial aversion learning in rats conditioned in their home cage. COA, like CTA, tolerates long intervals between the ingested stimuli and the illness and is long-lasting. Transient inactivation of the IC during acquisition spared COA whereas it greatly impaired CTA. Similarly, blockade of protein synthesis in IC did not affect COA but prevented CTA consolidation. Moreover, IC inactivation before retrieval tests did not interfere with COA memory expression when performed either 2 days (recent memory) or 36 days after acquisition (remote memory). Similar IC inactivation impaired the retrieval of either recent or remote CTA memory. Altogether these findings indicate that the IC is not necessary for aversive odour memory whereas it is essential for acquisition, consolidation and retrieval of aversive taste memory. We propose that the chemosensory stimulations modulate IC recruitment during the formation and the retrieval of food aversive memory.     

Role of cortical cannabinoid CB1 receptor in conditioned taste aversion memory

The brain endocannabinoid system has been shown to play a role in memory, though the extent to which this role generalizes over different types and processes of memory is not yet determined. Here we show that the cannabinoid receptor 1 (CB1) plays differential roles in acquisition, extinction and reconsolidation of conditioned taste aversion (CTA) memory in the rat insular cortex, which contains the taste cortex. Activation of the CB1 receptor in the insular cortex inhibits acquisition and reconsolidation but not extinction, whereas blockade of the CB1 receptor promotes memory and blocks extinction of CTA, while having no apparent effect on reconsolidation. The CB1 ligands used in this study were incapable of substituting the unconditioned stimulus in CTA training. All in all, the data raise the possibility that the state of activity of the CB1 receptor in the insular cortex contributes to the encoding of hedonic valence that enters into association with taste items.     

Nucleus reuniens of the thalamus controls fear memory intensity,specificity and long‐term maintenance during consolidation

The thalamic nucleus reuniens (NR) has been shown to support bidirectional medial prefrontal cortex‐hippocampus communication and synchronization relevant for cognitive processing. Using non‐selective or prolonged inactivation of the NR, previous studies reported its activity positively modulates aversive memory consolidation. Here we examined the NR's role in consolidating contextual fear memories with varied strength, at both recent and more remote time points, using muscimol‐induced temporary inactivation in rats. Results indicate the NR negatively modulates fear memory intensity, specificity, and long‐term maintenance. The more intense, generalized, and enduring fear memory induced by NR inactivation during consolidation was less prone to behavioral suppression by extinction or reconsolidation disruption induced by clonidine, an alpha‐2 adrenergic receptor agonist. Lastly, we used immunohistochemistry for Arc protein, which is involved in synaptic modifications underlying memory consolidation, to investigate whether treatment condition and/or conditioning status could change its levels not only in the NR, but also in the hippocampus (dorsal and ventral CA1 subregions) and the medial prefrontal cortex (anterior cingulate, prelimbic and infralimbic subregions). Results indicate a significant imbalance in the number of Arc‐expressing neurons in the brain areas investigated in muscimol fear conditioned animals when compared with controls. Collectively, present results provide convergent evidence for the NR's role as a hub regulating quantitative and qualitative aspects of a contextual fear memory during its consolidation that seem to influence the subsequent susceptibility to experimental interventions aiming at attenuating its expression. They also indicate the selectivity and duration of a given inactivation approach may influence its outcomes.     

Inhibition of protein kinase A activity during conditioned taste aversion retrieval: interference with extinction or reconsolidation of a memory?

The involvement of the cAMP-dependent protein kinase A (PKA) signaling pathway in protein synthesis-dependent memory consolidation has been supported by studies of fear conditioning and conditioned taste aversion (CTA). The present experiment examined whether inhibition of PKA activity at the time of memory retrieval impedes or promotes subsequent extinction. When Rp-cAMPS was infused into the amygdala at the time of CTA testing (retrieval), extinction was accelerated. Results confirm recent findings that stored memories become more labile when they are retrieved and extend these findings to CTA memories.     

Extinction is not a sufficient condition to prevent fear memories from undergoing reconsolidation in the basolateral amygdala

Consolidated memories when reactivated may return to a state that requires protein synthesis in order to be restabilized (reconsolidation). It has been shown in a variety of systems that if reactivation induces significant extinction then extinction is the protein synthesis dependent memory state, rather than reconsolidation. Thus, extinction consolidation may prevent the memory from undergoing reconsolidation. We investigated whether such an interaction also exists between extinction and reconsolidation of fear memories within the amygdala, by using a within subjects experimental design. We found that inhibition of protein synthesis in the basolateral amygdala (BLA) impaired reconsolidation for both the briefly reactivated and extinguished fear memories suggesting that extinction is not a sufficient condition to prevent induction of reconsolidation in the amygdala. These findings demonstrate that extinction consolidation does not always interact with reconsolidation. Therefore, under these conditions, extinction is not a boundary condition on reconsolidation of fear memories in the basolateral amygdala.     

Establishing aversive, but not safe, taste memories requires lateralized pontine-cortical connections

Aversive and safe taste memory processing is dramatically disrupted by bilateral lesions of the pontine parabrachial nucleus (PBN). To determine how such lesions affect patterns of neuronal activation in forebrain, lesions were combined with assessment of cFos-like immunoreactivity (FLI) in insular cortex (IC) and amygdala after conditioned taste aversion (CTA) training. Increases in FLI in amygdala and IC, which are normally seen following novel (versus familiar) CS-US pairing, were eliminated after PBN lesions. This suggests that PBN lesions prevent transmission of critical CS and US information to forebrain regions for the processing of both aversive and safe taste memories. Unilateral asymmetrical lesions of PBN and IC blocked CTA acquisition as well as normal patterns of FLI in amygdala after novel CS-US pairing, an effect not seen when unilateral lesions were confined to a single hemisphere. The crossed-disconnection experiments provide compelling evidence that functional interactions between PBN and IC are required for CTA acquisition, but not for safe taste memory formation and retrieval. The dissociation between effects of the different types of lesions on safe and aversive taste memories supports emerging evidence that the neural underpinnings of the two types of taste learning differ.     

Basolateral amygdala glutamatergic activation enhances taste aversion through NMDA receptor activation in the insular cortex

In conditioned taste aversion (CTA), a subject learns to associate a novel taste with visceral malaise. Brainstem, limbic and neocortical structures have been implicated in CTA memory formation. Nevertheless, the role of interactions between forebrain structures during these processes is still unknown. The present experiment was aimed at investigating the possible interaction between the basolateral nucleus of the amygdala (BLA) and the insular cortex (IC) during CTA memory formation. Injection of a low dose of lithium chloride (30 mg/kg, i.p.) 30 min after novel taste consumption (saccharin 0.1%) induces a weak CTA. Unilateral BLA injection of glutamate (2 microg in 0.5 microL) just before low lithium induces a stronger CTA. Unilateral injection of an N-methyl-d-aspartate (NMDA) receptor antagonist (AP5, 5 microg in 0.5 microL) in IC has no effect. However, AP5 treatment in IC at the same time or 1 h after the ipsilateral BLA injection reverses the glutamate-induced CTA enhancement. Injection of AP5 in IC 3 h after BLA injection does not interfere with the glutamate effect. Moreover, the CTA-enhancing effect of glutamate was also blocked by contralateral IC injection of AP5 at the same time. These results provide strong evidence that NMDA receptor activation in the IC is essential to enable CTA enhancement induced by glutamate infusion in the BLA during a limited time period that extends to 1 but not to 3 hours. These findings indicate that BLA-IC interactions regulate the strength of CTA. The bilateral nature of these amygdalo-cortical interactions is discussed.     

Dissecting mechanisms of reconsolidation: octopamine reveals differences between appetitive and aversive memories in the crab Chasmagnathus

Ample evidence suggests that, when reactivated by a reminder, a consolidated memory may return to a labile state and needs to be stabilized again in order to persist, a process known as reconsolidation. In a previous study, performed in the crab Chasmagnathus, we found a dual role for the biogenic amine octopamine (OA) during memory consolidation. On the one hand, it was necessary for appetitive memory formation and, on the other, it had a deleterious effect on aversive memory consolidation. Thus, OA could be a good candidate to dissect the neurochemical mechanisms of appetitive and aversive reconsolidation. Here, we initially characterized the reconsolidation of an appetitive memory. Then, we compared appetitive reconsolidation with its aversive counterpart regarding the implication of OA in these processes, and contrasted them with previous findings obtained in the consolidation phase. Our results demonstrate that appetitive reconsolidation takes place when animals are re-exposed to the training context, as shown by the amnesic effect of cycloheximide when applied before the reminder. In addition, the no-reinforcement during the reminder is a necessary condition for appetitive reconsolidation to occur. Remarkably, appetitive reconsolidation is neither impaired by OA receptor antagonists nor facilitated by exogenous OA, whereas aversive reconsolidation can be interfered with by OA administration. Thus, our results indicate that appetitive reconsolidation does not involve OA signaling, while aversive reconsolidation is negatively modulated by OA. All in all, these results could constitute a step towards the identification of particular features of appetitive and aversive reconsolidation.     

Brain sites involved in fear memory reconsolidation and extinction of rodents

Fear memory is a motivational system essential for organisms survival having a central role in organization of defensive behaviors to threat. In the last years there has been a growing interest on conditioned fear memory reconsolidation and extinction, two specific phases of memorization process, both induced by memory retrieval. Understanding the mechanisms underlying these two mnemonic processes may allow to work out therapeutic interventions for treatment of human fear and anxiety disorders, such as specific phobias and post-traumatic stress disorder. Based on the use of one-trial conditioning paradigms, which allow to follow the evolution of a mnemonic trace in its various phases, the present paper has attempted to reorganize the current literature relative to the rodents highlighting both the role of several brain structures in conditioned fear memory reconsolidation and extinction and the selective cellular processes involved. A crucial role seems to be play by medial prefrontal cortex, in particular by prelimbic and infralimbic cortices, and by distinct connections between them and the amygdala, hippocampus and entorhinal cortex.     

Differential involvement of cortical muscarinic and NMDA receptors in short- and long-term taste aversion memory

In conditioned taste aversion, an animal avoids a taste previously associated with toxic effects, and this aversive memory formation requires an intact insular cortex. In this paper, we investigated the possible differential involvement of cholinergic and glutamatergic receptors in the insular cortex in short-term memory (STM) and long-term memory (LTM) of taste aversion in rats. Taste aversion was induced by intraperitoneal administration of lithium chloride (a malaise-inducing drug) 15 min after experience with an unfamiliar taste. In order to test STM and LTM of taste aversion, taste stimulus was again presented 4 h and 72 h after lithium injection, respectively. During the acquisition, microinjection of the muscarinic antagonist, scopolamine, in the insular cortex before, but not after, the presentation of the new taste, abolished STM as well as LTM. Blockade of the NMDA receptor, in the insular cortex, by AP5 before, but not after, the presentation of the taste stimulus, impaired LTM but left STM intact. Moreover, when injected 1 h after malaise induction (i.e., during taste-illness association), AP5 disrupted both STM and LTM. These results suggest that activation of muscarinic receptors in the insular cortex is involved in the acquisition of taste memory, whereas NMDA receptors participate in taste memory consolidation. These data demonstrate that different neurochemical mechanisms subserve different memory phases. NMDA receptors are also probably involved in processing the visceral input, thus allowing subsequent taste-illness association. This indicates that in the same cortical area the same neurotransmitter system can be involved in distinct processes: taste memory consolidation vs. taste-illness association.     

Active Transition of Fear Memory Phase from Reconsolidation to Extinction through ERK-Mediated Prevention of Reconsolidation

The retrieval of fear memory induces two opposite memory process, i.e., reconsolidation and extinction. Brief retrieval induces reconsolidation to maintain or enhance fear memory, while prolonged retrieval extinguishes this memory. Although the mechanisms of reconsolidation and extinction have been investigated, it remains unknown how fear memory phases are switched from reconsolidation to extinction during memory retrieval. Here, we show that an extracellular signal-regulated kinase (ERK)-dependent memory transition process after retrieval regulates the switch of memory phases from reconsolidation to extinction by preventing induction of reconsolidation in an inhibitory avoidance (IA) task in male mice. First, the transition memory phase, which cancels the induction of reconsolidation, but is insufficient for the acquisition of extinction, was identified after reconsolidation, but before extinction phases. Second, the reconsolidation, transition, and extinction phases after memory retrieval showed distinct molecular and cellular signatures through cAMP responsive element binding protein (CREB) and ERK phosphorylation in the amygdala, hippocampus, and medial prefrontal cortex (mPFC). The reconsolidation phase showed increased CREB phosphorylation, while the extinction phase displayed several neural populations with various combinations of CREB and/or ERK phosphorylation, in these brain regions. Interestingly, the three memory phases, including the transition phase, showed transient ERK activation immediately after retrieval. Most importantly, the blockade of ERK in the amygdala, hippocampus, or mPFC at the transition memory phase disinhibited reconsolidation-induced enhancement of IA memory. These observations suggest that the ERK-signaling pathway actively regulates the transition of memory phase from reconsolidation to extinction and this process functions as a switch that cancels reconsolidation of fear memory.SIGNIFICANCE STATEMENT Retrieval of fear memory induces two opposite memory process; reconsolidation and extinction. Reconsolidation maintains/enhances fear memory, while extinction weakens fear memory. It remains unknown how memory phases are switched from reconsolidation to extinction during retrieval. Here, we identified an active memory transition process functioning as a switch that inhibits reconsolidation. This memory transition phase showed a transient increase of extracellular signal-regulated kinase (ERK) phosphorylation in the amygdala, hippocampus and medial prefrontal cortex (mPFC). Interestingly, inhibition of ERK in these regions at the transition phase disinhibited the reconsolidation-mediated enhancement of inhibitory avoidance (IA) memory. These findings suggest that the transition memory process actively regulates the switch of fear memory phases of fear memory by preventing induction of reconsolidation through the activation of the ERK-signaling pathway.     

Dynamic processing of taste aversion extinction in the brain

While substantial advances have been made in discovering how the brain learns and remembers, less is known about how the brain discards information, reorganizes information, or both. These topics are not only relevant to normal brain functioning but also speak to pathologies in which painful memories do not wane but are evoked time and again (e.g., post-traumatic stress disorder; PTSD). Here, we measured brain activity (as indicated by the regional expression of c-Fos protein) in rats during acquisition and throughout extinction of a conditioned taste aversion (CTA). We compared that brain activity with animals that had intact CTA memories or those that experienced an explicitly unpaired (EU) conditioned stimulus (CS; saccharin, SAC) and unconditioned stimulus (US; lithium chloride, LiCl). The data show a dynamic and nonuniform pattern of c-Fos protein expression in brain nuclei known to mediate gustation and CTAs. In particular, brainstem nuclei (e.g., nucleus of the solitary tract; NTS) and the basolateral nucleus of the amygdala (BLA) are active early as CTAs are formed and as extinction of the learned response begins. Later in the extinction process, the BLA reduces c-Fos expression relative to nonextinguished controls. Finally, as almost full reacceptance of the taste is achieved, the gustatory neocortex (GNC) expresses enhanced levels of c-Fos protein. Thus, extinction of a CTA is not represented by a simple reversal of the c-Fos activity evoked by CTA conditioning. Rather, the data demonstrate that extinction of conditioned responses is a dynamic process in which the activity levels of particular nuclei along the brain's taste pathway change depending on the extent to which the conditioned response has been extinguished.     

Reactivation with a simple exposure to the experimental environment is sufficient to induce reconsolidation requiring protein synthesis in the hippocampal CA3 region in mice

Our understanding of the memory reconsolidation process is at an earlier stage than that of consolidation. For example, it is unclear if, as for memory consolidation, reconsolidation of a memory trace necessitates protein synthesis. In fact, conflicting results appear in the literature and this discrepancy may be due to differences in the experimental reactivation procedure. Here, we addressed the question of whether protein synthesis in the CA3 hippocampal region is crucial in memory consolidation and reconsolidation of allocentric knowledge after reactivation in different experimental conditions in the Morris water maze. We showed (1) that an injection of the protein synthesis inhibitor anisomycin in the CA3 region during consolidation or after a single reactivation trial disrupted performance and (2) that protein synthesis is required even after a simple contextual reactivation without any learning trial and independently of the presence of the reinforcement. This work demonstrates that a simple exposure to the spatial environment is sufficient to reactivate the memory trace, to make it labile, and that reconsolidation of this trace requires de novo protein synthesis.     

Reconsolidation of fresh, remote, and extinguished fear memory in Medaka: old fears don't die

Long-term fear memory in the medaka fish (Oryzias latipes) regains transient sensitivity to a consolidation blocker immediately after memory reactivation in retrieval ('reconsolidation'). Here we show that reconsolidation occurs in fresh long-term memories but not in remote memories, and that the apparent amnesia induced by blockade of reconsolidation can be reinstated by an unpaired reinforcer, a procedure that has no effect on amnesia induced by blockade of consolidation. Extinction memory also undergoes post-reactivation reconsolidation, the blockade of which exposes the previously acquired fear. Hence in medaka, the process manifested in reconsolidation seems itself to consolidate; moreover, even when the post-reactivation application of the consolidation blocker is still able to disrupt the memory, the conditioned fear does not seem to go away permanently.     

Protein synthesis inhibition in the basolateral amygdala following retrieval does not impair expression of morphine-associated conditioned place preference

Conditioned place preference is an animal model used to evaluate the affective properties of natural rewards and drugs of abuse. This animal model is a kind of classical conditioning that depends on learning and memory. The basolateral amygdala (BLA) plays an important role in the consolidation and extinction of memory for this task. However, there is a lack of evidence demonstrating protein synthesis dependent reconsolidation following retrieval in conditioned animals. In other words, is it possible to observe morphine-associated place preference if recall of this preference is disrupted? Accordingly, we investigated this hypothesis by BLA infusion of protein synthesis inhibitor, anisomycin, immediately after retrieval (test) in conditioned place preference paradigm. In the first experiment, the conditioned animals were exposed to the two sides of the apparatus for 15 min in a drug-free state during retrieval. In the second experiment, the animals received an injection of morphine (7.5 mg/kg, i.p.) and immediately after, they were exposed to the two sides of the apparatus for 15 min. Finally in the third experiment, after habituation and training in the conditioned place preference task, the animals received an injection of the unconditioned stimulus (morphine, i.p.; 7.5 mg/kg) followed by confinement for 10 min in the morphine-paired compartment (conditioned stimulus) during memory retrieval. For the three experiments the animals were subsequently exposed in a free-drug state to the two sides of the apparatus for the retest. Our results show that the protein synthesis inhibition in all of these experimental designs had no effect on conditioned place preference memory under conditions that would initiate reconsolidation, suggesting that if reconsolidation of a conditioned place preference task exists it is not mediated by protein synthesis in basolateral amygdala. The effect of anisomycin on consolidation of contextual fear conditioning was also investigated as a positive control to assure that the negative results were not due to methodological problems. Using the same dose of anisomycin (62.5 microg/1 microl) in morphine-associated place preference procedures, we have found that this anisomycin dose blocks the consolidation of contextual fear memory, ruling out the possibility that these negative results can be attributed to methodological problem of some sort.     

Hand-eye coordination relies on extra-retinal signals: evidence from reactive saccade adaptation

Brain-derived neurotrophic factor (BDNF) has emerged as an important molecular mediator of synaptic plasticity. Our previous studies on the insular cortex (IC), a region of the temporal cortex implicated in the acquisition and storage of conditioned taste aversion (CTA), have demonstrated that the intracortical microinfusion of BDNF induces a lasting potentiation of synaptic efficacy in the projection from the basolateral nucleus of the amygdala (Bla) to the IC of adult rats in vivo. Recently, we have found that intracortical microinfusion of BDNF previous to CTA training modifies the retention of this task. In this work, we present experimental data showing that BDNF effects on CTA retention are dependent on both the activation of mitogen-activated protein kinases (MAPK) and phosphatidylinositol-3-kinase (PI-3K) at the insular cortex. Our results are evidence of the crucial role of both pathways in the modification of the CTA trace of memory caused by BDNF at a neocortical area.     

Impairment of aversive memory reconsolidation by localized intracranial electrical stimulation

Reconsolidation of long-term memory is blocked in animal models by macromolecular synthesis inhibitors, resulting in item-specific post-retrieval amnesia. The induction of such amnesia could ameliorate traumatic memories and phobias. However, this pharmacological approach is of limited value in humans because of toxicity. Here we report that reconsolidation of conditioned taste aversion in the rat is impaired by localized intracranial electrical stimulation. Lasting impairment was obtained only when stimulation was applied during memory reactivation and only to the dysgranular insular cortex bilaterally, which subserves the memory, but not to adjacent brain sites. The ability to learn a new association was not affected. The same method blocked new memory consolidation, but produced anterograde amnesia, reminiscent of the known effect of non-localized electroconvulsive therapy. Our results suggest that localized electrical microstimulation, such as produced by deep-brain stimulation or deep transcranial magnetic stimulation, could be used to impair long-term memory if applied during memory reactivation, and could lead to the development of a novel treatment for intractable post-traumatic stress disorder.     

A molecular switch for translational control in taste memory consolidation

In a variety of species memory consolidation following different learning paradigms has been shown to be dependent on protein synthesis. However, it is not known whether modulation of protein synthesis is a critical component of the consolidation process, nor is the identity of any protein(s) subject to translational regulation, known. We report here that phosphorylation of eukaryotic elongation factor-2 (eEF2), an indicator for translational elongation attenuation, is correlated with input that produces taste memory consolidation in the relevant cortex of rat. The temporal pattern of eEF2 phosphorylation is similar to extra-cellular regulated kinase 2 (ERK2) activation and S6K1 phosphorylation, which are known to stimulate translation initiation. In addition, increased eEF2 phosphorylation and increased alphaCaMKII expression is detected in a synaptoneurosomal fraction made from taste cortex following memory consolidation. These results suggest that increased initiation rate together with decreased elongation rate, during memory consolidation, shift the rate-limiting step of protein synthesis, to produce a local switch-like effect in the expression of neuronal proteins.     

The amygdalar circuit that acquires taste aversion memory differs from the circuit that extinguishes it

Experimental extinction is the decline in the frequency or intensity of a conditioned behaviour resulting from repetitive performance of the behaviour in the absence of the unconditioned stimulus or reinforcer ( Pavlov, 1927 ). Ample behavioural evidence indicates that experimental extinction does not reflect unlearning of the original trace, but rather a relearning process, in which the new association of the conditioned stimulus with the absence of the original reinforcer comes to control behaviour ( Rescorla, 1996 ). If experimental extinction is indeed learning rather than forgetting, are the neuronal circuits that subserve learning and extinction identical? We address this question by double dissociation analysis of the role of the central (CeA) and the basolateral (BLA) nuclei of the rat's amygdala in the acquisition and extinction, respectively, of conditioned taste aversion (CTA). Whereas local blockade of protein synthesis or β‐adrenergic receptors in the CeA blocks acquisition but not extinction of CTA, a similar intervention in the BLA blocks extinction but not acquisition. Hence, the amygdalar circuit that acquires taste aversion memory differs functionally from the circuit that extinguishes it.