Context modulation of memory for fear extinction in humans

Distinct memories are formed during fear conditioning and subsequent extinction. In animals, the expression of the latter is gated by the context. The recall of extinction memory after a long delay, and the contextual modulation thereof, has not been directly tested in humans. Mentally healthy volunteers underwent a 2-day fear conditioning and extinction protocol that examined the recall of extinction memory and its relationship to context. Conditioned stimuli were paired with an aversive electric shock in one visual context and extinguished in a different context. Extinction recall and renewal were examined 24 h after training. We found that skin conductance responses were small when the conditioned stimulus was presented in the extinction context, but responses were renewed when the conditioned stimulus was presented in the conditioning context. This finding demonstrates context dependency of extinction recall in humans.     

Peripheral vasopressin accelerates extinction of conditioned taste avoidance

Both peripheral and central administration of vasopressin improves retention and delays extinction when given before or after acquisition of shock avoidance learning. For conditioned taste avoidance, however, vasopressin prolongs extinction when injected peripherally before acquisition tests and accelerates extinction when infused intracerebroventricularly after acquisition. The following experiments were designed to determine whether this inconsistency is based on the route of administration or timing of vasopressin treatment. Because acquisition of conditioned taste avoidance is strengthened when an agent that is capable of inducing avoidance is administered after LiCl injection, it was determined in experiment 1 that a 6 microg/kg dose of vasopressin did not induce conditioned taste avoidance when administered 50 min after consumption of a sucrose solution. In experiment 2, it was determined that this dose of vasopressin accelerated extinction of a LiCl-induced conditioned taste avoidance when given 50 min after LiCl injection. These results suggest that the inconsistency is not based on route of administration. In experiment 3, it was determined that there was a tendency for animals to show prolonged extinction when vasopressin was administered 20 min before access to a sucrose solution. All of the results taken together suggest that the differential effects of vasopressin on extinction are due to the timing of administration. It was suggested that vasopressin accelerates extinction when given after acquisition by reducing the effectiveness of LiCl and it prolongs extinction when given before acquisition by altering neural responsiveness in areas mediating conditioned taste avoidance.     

Experience with activity based anorexia enhances conditioned taste aversion learning in rats

Activity based anorexia (ABA) is a model that mimics the self-starvation and hyperactivity features of anorexia nervosa (AN). This study investigated whether a history of ABA will enhance food avoidance learning and retard its extinction in female rats. We compared the acquisition and extinction of a conditioned taste aversion (CTA) in naive (ad lib with no access to RW), ABA, and pair-fed to the food intake of ABA (with access to a locked RW) female Sprague-Dawley rats. The CTA conditioning was conducted after the ABA and pair-fed rats had recovered to their pre-food restriction body weights. For the CTA learning, 0.3 M sucrose consumption was followed by low doses LiCl (0.009 M or 0.018 M at 1.33 ml/100 g of body weight, IP) injection. The results revealed that the ABA rats acquired an aversion to sucrose significantly sooner than the naive controls. Furthermore, they completely avoided sucrose while the naive and pair-fed controls still sampled it by the end of 10 conditioning trials. When extinction was assessed by 1-bottle and 2-bottle tests, the ABA rats extinguished more slowly than the controls. However, the differences in sucrose aversion extinction between the ABA and control rats were only significant in the 1-bottle test. These data suggest that experience with AN-like behaviors results in an acquired aversion to a preferred food sooner and a longer retention of the negative food associations. These findings have implications for understanding the persistence of aberrant eating behaviors in eating disorders.     

c-Fos Induction in the Rat Nucleus of the Solitary Tract by Intraoral Quinine Infusion Depends on Prior Contingent Pairing of Quinine and Lithium Chloride

Intraoral infusions of sucrose or saccharin induce c-Fos-like immunoreactivity (c-FLI) in the intermediate nucleus of the solitary tract (iNTS) of rats after acquisition of a conditioned taste aversion (CTA). The induction of c-FLI in the iNTS may be a consequence of the shift in behavioral response from ingestive to aversive behaviors that characterize acquisition and expression of a CTA. To test this hypothesis, rats were intraorally infused with 0.3 mM quinine sulfate, an aversive taste, 1. prior to conditioning, 2. after 3 noncontingent (unpaired) infusions of quinine and toxic lithium chloride (LiCl) injections, 3. after conditioning with 3 contingent pairings of quinine and LiCl, and 4. after extinction with repeated unpaired infusions of quinine. Intraoral infusions of quinine induced c-FLI in the iNTS only after acquisition of a CTA against quinine; quinine failed to induce c-FLI in the iNTS of unconditioned, noncontingently treated, or extinguished rats. The pattern of c-FLI in the iNTS induced by expression of a CTA against quinine was quantitatively and anatomically similar to that elicited by sucrose in rats expressing a CTA against sucrose. We conclude that aversive responses per se are not sufficient to induce c-FLI in the iNTS. Furthermore, contingent pairing of quinine and LiCl does not cause a shift in behavioral response from palatable, ingestive behaviors to aversive behaviors as in acquisition of a CTA against sucrose. Thus, we also conclude that a shift in behavior from ingestive to aversive responses is not required for increased c-FLI expression in the iNTS during CTA expression. Therefore, the induction of c-FLI in the iNTS during expression of a CTA may be correlated with neuronal processes specific to acquisition and expression of a CTA.     

Effects of midazolam on acquisition and extinction of conditioned taste aversion memory in rats

Some intravenous anesthetic agents such as midazolam are known to induce anterograde and retrograde amnesia. We analyzed the effect of midazolam by the conditioned taste aversion (CTA) acquisition and retention. After the rats were offered 0.1% sodium saccharin (Sac) as conditioned stimulus (CS), an intraperitoneal (i.p.) injection of several concentrations (5–30 mg/kg) of midazolam was followed by an i.p. injection of 0.15 M LiCl (2% of body weight) as unconditioned stimulus (US). The rats, which acquired CTA by every CS–US paradigm, strongly avoided Sac on the 1st test day after conditioning and maintained the avoidance for 3 days. We have already reported that Sac intake abruptly increased on the 2nd test day and the almost complete extinction occurred on the 3rd test day after conditioning by injection of subhypnotic dose of propofol before LiCl-injection. In contrast, we found that subhypnotic dose of midazolam suppressed not only CTA acquisition, but also CTA retention. On the other hand, an α2-adrenergic blocker, yohimbin (1 mg/kg) suppressed only the CTA retention. These results suggest that the subhypnotic doses of midazolam firstly affect the acquisition mechanism of the CTA memory (CTAM), resulting the suppression of the retention of CTAM.     

High doses of vasopressin delay the onset of extinction and strengthen acquisition of LiCl-induced conditioned taste avoidance

When low doses of vasopressin are given 50 min after pairing sucrose consumption with a high dose of LiCl, extinction of the LiCl-induced conditioned taste avoidance is accelerated. These low doses of vasopressin do not themselves induce conditioned taste avoidance when paired with sucrose consumption. Predicated on previous studies administering two avoidance-inducing agents after sucrose consumption, studies were designed to determine whether high doses of vasopressin capable of inducing conditioned taste avoidance would (1) delay rather than accelerate extinction of a conditioned taste avoidance induced by a high dose of LiCl and (2) strengthen acquisition of a conditioned taste avoidance induced by a low dose of LiCl. The results of three studies showed that doses of 9 and 18 microg/kg of vasopressin induced a conditioned taste avoidance when injected 50 min after sucrose consumption, delayed the onset of extinction when injected 50 min after pairing sucrose consumption with a high dose of LiCl, and strengthened acquisition of a conditioned taste avoidance when injected 50 min after pairing sucrose consumption with a low dose of LiCl. Taken together, these data suggest that the delay in onset of extinction is due to a strengthening of acquisition. It has been suggested that vasopressin is a mnemonic neuropeptide that delays extinction of learned tasks. However, for conditioned taste avoidance, the evidence for the effects of low doses of vasopressin on extinction do not support this hypothesis and the evidence for high doses of vasopressin can be accounted for by the avoidance-inducing properties of vasopressin.     

Cortical involvement in acquisition and extinction of trace eyeblink conditioning

Previous studies have implicated 2 cortical regions interconnected with the hippocampal formation, the retrosplenial cortex (RSC) and the medial prefrontal cortex (mPFC), as loci important for the acquisition of hippocampally dependent trace eyeblink conditioning. These loci have also been proposed to serve as long-term storage sites of task critical information. This study used lesions made prior to training to investigate the roles of the RSC, as well as the caudal and rostral subdivisions of the mPFC, in the acquisition and subsequent extinction of trace eyeblink conditioning in the rabbit. The caudal mPFC and rostral mPFC were shown to be critical for acquisition and extinction of the conditioned reflex, respectively. The data indicate that the RSC is not critical for acquisition or extinction of the trace conditioned reflex.     

Effects of psilocybin on hippocampal neurogenesis and extinction of trace fear conditioning

Drugs that modulate serotonin (5-HT) synaptic concentrations impact neurogenesis and hippocampal (HPC)-dependent learning. The primary objective is to determine the extent to which psilocybin (PSOP) modulates neurogenesis and thereby affects acquisition and extinction of HPC-dependent trace fear conditioning. PSOP, the 5-HT_2A agonist 25I-NBMeO and the 5-HT_2A/C antagonist ketanserin were administered via an acute intraperitoneal injection to mice. Trace fear conditioning was measured as the amount of time spent immobile in the presence of the conditioned stimulus (CS, auditory tone), trace (silent interval) and post-trace interval over 10 trials. Extinction was determined by the number of trials required to resume mobility during CS, trace and post-trace when the shock was not delivered. Neurogenesis was determined by unbiased counts of cells in the dentate gyrus of the HPC birth-dated with BrdU co-expressing a neuronal marker. Mice treated with a range of doses of PSOP acquired a robust conditioned fear response. Mice injected with low doses of PSOP extinguished cued fear conditioning significantly more rapidly than high-dose PSOP or saline-treated mice. Injection of PSOP, 25I-NBMeO or ketanserin resulted in significant dose-dependent decreases in number of newborn neurons in hippocampus. At the low doses of PSOP that enhanced extinction, neurogenesis was not decreased, but rather tended toward an increase. Extinction of “fear conditioning” may be mediated by actions of the drugs at sites other than hippocampus such as the amygdala, which is known to mediate the perception of fear. Another caveat is that PSOP is not purely selective for 5-HT_2A receptors. PSOP facilitates extinction of the classically conditioned fear response, and this, and similar agents, should be explored as potential treatments for post-traumatic stress disorder and related conditions.     

Timing of extinction relative to acquisition: a parametric analysis of fear extinction in humans

Fear extinction is a reduction in conditioned fear following repeated exposure to the feared cue in the absence of any aversive event. Extinguished fear often reappears after extinction through spontaneous recovery. Animal studies suggest that spontaneous recovery can be abolished if extinction occurs within minutes of acquisition. However, a limited number of human extinction studies have shown that short interval extinction does not prevent the return of fear. For this reason, we performed an in-depth parametric analysis of human fear extinction using fear-potentiated startle. Using separate single-cue and differential conditioning paradigms, participants were fear conditioned and then underwent extinction either 10 min (Immediate) or 72 hr (Delayed) later. Testing for spontaneous recovery occurred 96 hr after acquisition. In the single cue paradigm, the Immediate and Delayed groups exhibited differences in context, but not fear, conditioning. With differential conditioning, there were no differences in context conditioning and the Immediate group displayed less spontaneous recovery. Thus, the results remain inconclusive regarding spontaneous recovery and the timing of extinction and are discussed in terms of performing translational studies of fear in humans.     

The effect of androgen on the retention of extinction memory after conditioned taste aversion in mice

Conditioned taste aversion (CTA) induced by the application of a novel taste such as sodium saccharin (Sac) as the conditioned stimulus (CS) and a malaise-inducing agent as the unconditioned stimulus (US), results in acquisition of CTA memory to Sac. In contrast, CTA is extinguished by repeated presentations of the CS without the US, resulting in acquisition of the extinction memory. We examined the effects of androgenic hormones on acquisition and retention of extinction memory in mice. We gonadectomized sexually immature mice and continuously administered androgens to these animals. After sexual maturation, the mice underwent a conditioning period followed by an extinction period. Retrieval tests revealed that the androgen-treated group showed significantly greater retention of extinction memory than the non-treated group 5 weeks later, whereas such significant difference was not observed in acquisition of extinction memory. These results demonstrate the enhancing effect of androgens on retention of extinction memory.     

Influence of ethanol withdrawal on fear memory: Effect of D-cycloserine

Animals made dependent via an ethanol (ETOH) -containing liquid diet (6% v/v) for 14 days were subjected to a contextual fear conditioning paradigm 3 days after the last consumption day. After conditioning, rats were subjected to four extinction trials by exposing the animals to the conditioned context and their freezing was evaluated for each trial. Immediately after the first extinction trial, animals were injected with D-cycloserine (DCS) 5 mg/kg i.p., a dose that did not influence the extinction in control rats. Spontaneous recovery of learned fear was tested seven days after the last extinction trial. The following day, animals were subjected to a reacquisition or a reinstatement procedure and their freezing responses evaluated 24 h later. The present study shows that: 1. discontinuation from chronic ETOH administration facilitated the formation of a new fear memory concomitant with a marked resistance to being extinguished, 2. administration of DCS (5 mg/kg) facilitated the extinction process only in ETOH withdrawn rats, 3. both reinstatement and reacquisition procedures restored the increased freezing in ETOH withdrawn animals after extinction, 4. DCS administered immediately after the first extinction trial prevented the increase in freezing following both reacquisition and reinstatement. The enhanced sensitivity to the facilitatory effect of DCS in ETOH withdrawn animals may be mediated by adaptive changes in N-methyl-D-aspartate (NMDA) receptor provoked by ETOH dependence.     

Conditioned and extinguished fear modulate functional corticocardiac coupling in humans

Although the conditioned cardiac fear response is an important index of psychophysiological fear processing, underlying neural mechanisms remain unclear. N = 22 participants underwent differential fear conditioning and extinction with face pictures as conditioned stimuli (CS) and loud noise bursts as aversive unconditioned stimulus (US) on Day 1 and a recall test 1 day later. We assessed ERPs, evoked heart period (HP), and time‐lagged within‐subject correlations of single‐trial EEG amplitude and HP as index for corticocardiac coupling in response to the CS. Fear‐conditioned stimuli (CS+) triggered cardiac deceleration during fear acquisition and recall. Meanwhile, only during Day 1 acquisition, CS+ evoked larger late positivities in the ERP than CS?. Most importantly, during Day 2 recall, stimulus‐evoked single‐trial EEG responses in the time window between 250 and 500 ms predicted the magnitude of cardiac fear responses 2 to 5 s later. This marker of corticocardiac coupling selectively emerged in response to not previously extinguished CS+ but was absent in response to CS? or previously extinguished CS+. The present results provide first evidence that fear conditioning and extinction modulate functional corticocardiac coupling in humans. Underlying mechanisms may involve subcortical structures enhancing corticocardiac transmission to facilitate processing of consolidated conditioned fear.     

Activation of basolateral amygdala corticotropin-releasing factor 1 receptors modulates the consolidation of contextual fear

The basolateral amygdala complex (BLA) and central amygdala nucleus (CeA) are involved in fear and anxiety. In addition, the BLA contains a high density of corticotropin-releasing factor 1 (CRF₁) receptors in comparison to the CeA. However, the role of BLA CRF₁ receptors in contextual fear conditioning is poorly understood. In the present study, we first demonstrated in rats that oral administration of DMP696, the selective CRF₁ receptor antagonist, had no significant effects on the acquisition of contextual fear but produced a subsequent impairment in contextual freezing suggesting a role of CRF₁ receptors in the fear memory consolidation process. In addition, oral administration of DMP696 significantly reduced phosphorylation of cyclic AMP response element-binding protein (pCREB) in the lateral and basolateral amygdala nuclei, but not in the CeA, during the post-fear conditioning period. We then demonstrated that bilateral microinjections of DMP696 into the BLA produced no significant effects on the acquisition of conditioned fear but reduced contextual freezing in a subsequent drug-free conditioned fear test. Importantly, bilateral microinjections of DMP696 into the BLA at 5 min or 3 h, but not 9 h, after exposure to contextual fear conditioning was also effective in reducing contextual freezing in the conditioned fear test. Finally, microinfusions of either DMP696 into the CeA or a specific corticotropin-releasing factor 2 receptor antagonist in the BLA were shown to have no major effects on disrupting either contextual fear conditioning or performance of contextual freezing in the drug-free conditioned fear test. Collectively, results implicate a role of BLA CRF₁ receptors in activating the fear memory consolidation process, which may involve BLA pCREB-induced synaptic plasticity.     

The role of different subregions of the basolateral amygdala in cue-induced reinstatement and extinction of food-seeking behavior

Reinstatement of previously extinguished instrumental responding for drug-related cues has been used as an animal model for relapse of drug abuse, and is disrupted by inactivation of the basolateral amygdala (BLA). However, the role that the BLA plays in reinstatement induced by cues associated with natural rewards is unclear. The present study assessed the effects of inactivation of different regions of the BLA in cue-induced reinstatement of food-seeking behavior and in the extinction of instrumental responding for food. In experiment 1, rats acquired a lever pressing response for food reward paired with a light/tone conditioned stimulus (CS). They were then subjected to extinction training, where both food and the CS were withheld. Reinstatement of extinguished responding was measured during response-contingent presentations of the CS alone. Following saline infusions into the caudal or rostral BLA, rats displayed a significant increase in lever pressing during reinstatement sessions. Inactivation of these subregions with bupivacaine did not attenuate responding for the CS in the absence of food delivery. In fact, inactivation of the caudal BLA potentiated responding relative to vehicle treatments. Analysis of within-session responding revealed that caudal BLA inactivation retarded extinction of lever pressing in response to the CS. In experiment 2, inactivation of the caudal BLA on the first or second day of extinction training significantly retarded the acquisition of extinction learning on the following day. These data indicate that that the caudal BLA may play a specific role in the extinction of appetitive conditioned responses, by monitoring changes in the reinforcing value of pavlovian conditioned stimuli linked to action-outcome associations once these associations have been formed. Moreover, these findings support a growing body of evidence indicating that separate neural circuits incorporating the BLA may play different roles in mediating reinstatement of reward-seeking behaviors induced by either drug or food related stimuli.     

Temporal and qualitative dynamics of conditioned taste aversion processing: combined generalization testing and licking microstructure analysis

The pattern of licking microstructure during various phases of a conditioned taste aversion (CTA) was evaluated. In Experiment 1, rats ingested lithium chloride (LiCl) for 3 trials and were then offered sodium chloride (NaCl) or sucrose on 3 trials. A CTA to LiCl developed and generalized to NaCl but not to sucrose. CTA intake suppression was characterized by reductions in burst size, average ingestion rate, and intraburst lick rate, and increases in brief pauses and burst counts. Compared with previous studies, LiCl licking shifted from a pattern initially matching that for normally accepted NaCl to one matching licking for normally avoided quinine hydrochloride by the end of the 1st acquisition trial. In Experiment 2, a novel paradigm was developed to show that rats expressed CTA generalization within 9 min of their first LiCl access. These results suggest that licking microstructure analysis can be used to assay changes in hedonic evaluation caused by treatments that produce aversive states.     

The effects of neurotoxic hippocampal lesions on two effects of context after fear extinction

Three conditioned suppression experiments with rats examined the role of the hippocampus in 2 effects of context after extinction. Reinstatement is the context-specific recovery of fear to an extinguished conditioned stimulus (CS) that occurs following independent presentations of the unconditioned stimulus (US), after extinction. Renewal is the recovery of fear when the CS is presented in the context in which it was conditioned, after extinction in a different context. Results indicated that neurotoxic lesions of the hippocampus, performed before conditioning, abolished reinstatement, which depends on context-US associations, but not renewal, which does not. This dissociation is not the result of differences in the recentness of context learning that ordinarily governs the 2 effects. The results suggest that the hippocampus is necessary for some, but not all, types of contextual learning.     

Network model of fear extinction and renewal functional pathways

The objective of this study was to examine the opposite behavior responses of conditioned fear extinction and renewal and how they are represented by network interactions between brain regions. This work is a continuation of a series of brain mapping studies of various inhibitory phenomena, including conditioned inhibition, blocking and extinction. A tone-footshock fear conditioning paradigm in rats was used, followed by extinction and testing in two different contexts. Fluorodeoxyglucose autoradiography was used to compare mean regional brain activity and interregional correlations resulting from the presentation of the extinguished tone in or out of the extinction context. A confirmatory structural equation model, constructed from a neural network proposed to underlie fear extinction, showed a reversal from negative regional interactions during extinction recall to positive interactions during fear renewal. Additionally, the magnitude of direct effects was different between groups, reflecting a change in the strength of the influences conveyed through those pathways. The results suggest that the extinguished tone encountered outside of the extinction context recruits auditory and limbic areas, which in turn influence the interactions of the infralimbic cortex with the amygdala and ventrolateral periaqueductal gray. Interestingly, the results also suggest that two independent pathways influence conditioned freezing: one from the central amygdaloid nucleus and the other from the infralimbic cortex directly to the ventrolateral periaqueductal gray.     

Opioid receptors regulate the extinction of Pavlovian fear conditioning

Rats received a single pairing of an auditory conditioned stimulus (CS) with a footshock unconditioned stimulus (US). The fear (freezing) that had accrued to the CS was then extinguished. Injection of naloxone prior to this extinction significantly impaired the development of extinction. This impairment was mediated by opioid receptors in the brain and was not observed when naloxone was injected after extinction training. Finally, an injection of naloxone on test failed to reinstate extinguished responding that had already accrued to the CS. These experiments show that opioid receptors regulate the development, but not the expression, of fear extinction and are discussed with reference to the roles of opioid receptors in US processing, memory, and appetitive motivation.     

Reversible inactivations of the cerebellum with muscimol prevent the acquisition and extinction of conditioned nictitating membrane responses in the rabbit

Lesions of the cerebellum severely impair the classically conditioned nictitating membrane response (NMR) in rabbits. Thus, the cerebellum is essential for the production of conditioned responses (CRs), either because it is actively involved in NMR conditioning or because damage to it causes motor or other general deficits. To distinguish between these alternatives, the cerebellum may be inactivated during training. Inactivation of the cerebellum during acquisition training might result in the absence of CRs on initial trials of subsequent training without the neuronal blockade. The blockade may have prevented learning but it may have produced other deficits that require time or further training to overcome. This problem can be addressed by inactivating the cerebellum during extinction training. If inactivation during extinction training results in the immediate production of CRs when training is resumed without the blockade, then it may be concluded that extinction learning was prevented by the blockade — the presence of CRs argues against any deficits not associated with learning. We used muscimol to inactivate the cerebellum and test its involvement in acquisition and extinction of NMR conditioning in the same subjects. We injected muscimol close to the interpositus nucleus of the cerebellum 1 h before each of four daily training sessions of delay conditioning. Almost no CRs were produced in these training sessions — there was little or no acquisition of NMR conditioning during cerebellar inactivation. The subjects were then trained for four daily sessions without injections of muscimol. There were no CRs on initial trials of the first session of retraining, but all subjects produced CRs by the end of this session. The subjects then received four daily sessions of extinction training with muscimol inactivation of the nuclei — no CRs were produced. Extinction training then continued for four daily sessions without muscimol inactivation. On the first of these sessions, all subjects immediately produced high levels of CRs. These responses then extinguished within and between sessions with characteristic beginning-of-session spontaneous recovery. There was little or no extinction of NMR conditioning during cerebellar inactivation. After inactivation, the muscimolinactivated subjects went on to acquire and extinguish NM responses at rates similar to those of appropriate controls. We conclude that cerebellar circuitry is essential for, and actively engaged in, both acquisition and extinction of this simple form of motor learning.     

The effect of yohimbine on the extinction of conditioned fear: a role for context

Six experiments with rat subjects examined the effect of yohimbine, an alpha-2 adrenergic autoreceptor antagonist, on the extinction of conditioned fear to a tone. Experiments 1 and 2 demonstrated that systemic administration of yohimbine (1.0 mg/kg) facilitated a long-term decrease in freezing after extinction, and this depended on pairing drug administration with extinction training. However, Experiments 3 and 4 demonstrated that yohimbine did not eradicate the original fear learning: Freezing was renewed when the tone was tested outside of the extinction context. Experiments 5 and 6 found that the contextually specific attenuation of fear produced by yohimbine transferred to another extinguished conditional stimulus (CS) and not to a nonextinguished CS. The results suggest that yohimbine, when administered in the presence of a neutral context, creates a form of inhibition in that context that allows that specific context to reduce fear of an extinguished CS.