Individuals use both passive and active defensive responses to environmental threats. Much is known about the neural circuits of passive defensive responses (e.g., freezing), but less is known about the substrates of active defensive responses (e.g., avoidance). We developed an active avoidance task in which rats learn to avoid a tone-signaled footshock by stepping onto a nearby platform. An advantage of this task is that freezing, which can interfere with avoidance, is reduced, thereby facilitating comparison of the effects of manipulations on avoidance versus freezing. After 10 d of avoidance training, rats were infused with muscimol to pharmacologically inactivate the prelimbic cortex (PL), infralimbic cortex (IL), ventral striatum (VS), or basolateral amygdala (BLA). Inactivating PL, VS, or BLA all impaired avoidance expression, but these areas differed with respect to freezing. Inactivating BLA decreased freezing consistent with loss of the tone-shock association, whereas inactivation of VS increased freezing consistent with loss of avoidance memory. Inactivation of PL had no effect on freezing. Inactivation of IL did not impair avoidance expression but did impair avoidance extinction. Our findings suggest that active avoidance is mediated by prefrontal-striatal circuits, which may be overactive in individuals suffering from trauma-related disorders.
Infusing brain-derived neurotrophic factor (BDNF) into the infralimbic (IL) prefrontal cortex is capable of inducing extinction. Little is known, however, about the circuits mediating BDNF effects on extinction or the extent to which extinction requires BDNF in IL. Using local pharmacological infusion of BDNF protein, or an antibody against BDNF, we found that BDNF in the IL, but not prelimbic (PL) prefrontal cortex, is both necessary and sufficient for fear extinction. Furthermore, we report that BDNF in IL can induce extinction of older fear memories (14 days) as well as recent fear memories (1 day). Using immunocytochemistry, we show that BDNF is increased in the ventral hippocampus (vHPC), but not IL or PL, following extinction training. Finally, we observed that infusing BDNF into the vHPC increased the firing rate of IL, but not PL neurons in fear conditioned rats. These findings indicate that an extinction-induced increase in BDNF within the vHPC enhances excitability in IL targets, thereby supporting extinction memories.
The acquisition and expression of conditioned fear depends on prefrontal-amygdala circuits. Auditory fear conditioning increases the tone responses of lateral amygdala neurons, but the increase is transient, lasting only a few hundred milliseconds after tone onset. It was recently reported that that the prelimbic (PL) prefrontal cortex transforms transient lateral amygdala input into a sustained PL output, which could drive fear responses via projections to the lateral division of basal amygdala (BL). To explore the possible mechanisms involved in this transformation, we developed a large-scale biophysical model of the BL-PL network, consisting of 850 conductance-based Hodgkin-Huxley-type cells, calcium-based learning, and neuromodulator effects. The model predicts that sustained firing in PL can be derived from BL-induced release of dopamine and norepinephrine that is maintained by PL-BL interconnections. These predictions were confirmed with physiological recordings from PL neurons during fear conditioning with the selective ?-blocker propranolol and by inactivation of BL with muscimol. Our model suggests that PL has a higher bandwidth than BL, due to PLs decreased internal inhibition and lower spiking thresholds. It also suggests that variations in specific microcircuits in the PL-BL interconnection can have a significant impact on the expression of fear, possibly explaining individual variability in fear responses. The human homolog of PL could thus be an effective target for anxiety disorders.
It is thought that discrete subregions of the medial prefrontal cortex (mPFC) regulate different aspects of appetitive behavior, however, physiological support for this hypothesis has been lacking. In the present study, we used multichannel single-unit recording to compare the response of neurons in the prelimbic (PL) and infralimbic (IL) subregions of the mPFC, in rats pressing a lever to obtain sucrose pellets on a variable interval schedule of reinforcement (VI-60). Approximately 25% of neurons in both structures exhibited prominent excitatory responses during rewarded, but not unrewarded, lever presses. The time courses of reward responses in PL and IL, however, were markedly different. Most PL neurons exhibited fast and transient responses at the delivery of sucrose pellets, whereas most IL neurons exhibited delayed and prolonged responses associated with the collection of earned sucrose pellets. We further examined the functional significance of reward responses in IL and PL with local pharmacological inactivation. IL inactivation significantly delayed the collection of earned sucrose pellets, whereas PL inactivation produced no discernible effects. These findings support the hypothesis that PL and IL signal distinct aspects of appetitive behavior, and suggest that IL signaling facilitates reward collection.
The most effective treatment for posttraumatic stress disorder (PTSD) is exposure therapy, which aims to facilitate extinction of conditioned fear. Recent evidence suggests that brain-derived neurotrophic factor (BDNF) facilitates extinction learning. This study assessed whether the Met-66 allele of BDNF, which results in lower activity-dependent secretion, predicts poor response to exposure therapy in PTSD.
Deep brain stimulation (DBS) of the ventral capsule/ventral striatum (VC/VS) reduces the symptoms of treatment-resistant obsessive compulsive disorder (OCD), and improves response to extinction-based therapies. We recently reported that DBS-like stimulation of a rat homologue of VC/VS, the dorsal-VS, reduced conditioned fear and enhanced extinction memory (Rodriguez-Romaguera et al., 2012). In contrast, DBS of the ventral-VS had the opposite effects. To examine possible mechanisms of these effects, we assessed the effects of VS DBS on the expression of the neural activity marker Fos and brain-derived neurotrophic factor (BDNF), a key mediator of extinction plasticity in prefrontal-amygdala circuits. Consistent with decreased fear expression, DBS of dorsal-VS increased Fos expression in prelimbic and infralimbic prefrontal cortices and in the lateral division of the central nucleus of amygdala, an area that inhibits amygdala output. Consistent with improved extinction memory, we found that DBS of dorsal-VS, but not ventral-VS, increased neuronal BDNF expression in prelimbic and infralimbic prefrontal cortices. These rodent findings are consistent with the idea that clinical DBS of VC/VS may augment fear extinction through an increase in BDNF expression.
Increasing evidence indicates that the medial prefrontal cortex (mPFC) and the amygdala mediate expression and extinction of conditioned fear, but few studies have examined the inputs to these structures. The dorsal part of the midline thalamus (dMT) contains structures such as the mediodorsal nucleus, paraventricular nucleus, and paratenial nucleus that project prominently to mPFC, as well as to basal (BA) and central (Ce) nuclei of the amygdala. Using temporary inactivation with GABA agonist muscimol, we found that dMT was necessary for retrieving auditory fear memory that was 24 h old, but not 2-8 h old. However, pre-training infusions did not impair fear acquisition or extinction. To determine the possible targets of dMT that might modulate fear retrieval, we combined dMT inactivation with Fos immunohistochemistry. Rats with inactivation-induced impairment in fear retrieval showed increased Fos in the lateral division of Ce (CeL), and decreased Fos in the medial division of Ce. No differences in Fos expression were observed in the mPFC or BA. We suggest that the projections from the paraventricular nucleus to CeL are involved in retrieval of well consolidated fear memories. This article is part of a Special Issue entitled Anxiety and Depression.
Decades of behavioral studies have confirmed that extinction does not erase classically conditioned fear memories. For this reason, research efforts have focused on the mechanisms underlying the development of extinction-induced inhibition within fear circuits. However, recent studies in rodents have uncovered mechanisms that stabilize and destabilize fear memories, opening the possibility that extinction might be used to erase fear memories. This symposium focuses on several of these new developments, which involve the timing of extinction training. Extinction-induced erasure of fear occurs in very young rats, but is lost with the development of perineuronal nets in the amygdala that render fear memories impervious to extinction. Moreover, extinction administered during the reconsolidation phase, when fear memory is destabilized, updates the fear association as safe, thereby preventing the return of fear, in both rats and humans. The use of modified extinction protocols to eliminate fear memories complements existing pharmacological strategies for strengthening extinction.
Current models of conditioned fear expression and extinction involve the basolateral amygdala (BLA), ventral medial prefrontal cortex (vmPFC), and the hippocampus (HPC). There is some disagreement with respect to the specific roles of these structures, perhaps due to subregional differences within each area. For example, growing evidence suggests that infralimbic (IL) and prelimbic (PL) subregions of vmPFC have opposite influences on fear expression. Moreover, it is the ventral HPC (vHPC), rather than the dorsal HPC, that projects to vmPFC and BLA. To help determine regional specificity, we used small doses of the GABA(A) agonist muscimol to selectively inactivate IL, PL, BLA, or vHPC in an auditory fear conditioning and extinction paradigm. Infusions were performed prior to extinction training, allowing us to assess the effects on both fear expression and subsequent extinction memory. Inactivation of IL had no effect on fear expression, but impaired the within-session acquisition of extinction as well as extinction memory. In contrast, inactivation of PL impaired fear expression, but had no effect on extinction memory. Inactivation of the BLA or vHPC impaired both fear expression and extinction memory. Post-extinction inactivations had no effect in any structure. We suggest a model in which amygdala-dependent fear expression is modulated by inputs from PL and vHPC, whereas extinction memory requires extinction-induced plasticity in IL, BLA, and/or vHPC.
Consolidation of fear extinction involves enhancement of N-methyl D aspartate (NMDA) receptor-dependent bursting in the infralimbic region (IL) of the medial prefrontal cortex (mPFC). Previous studies have shown that systemic blockade of metabotropic glutamate receptor type 5 (mGluR5) reduces bursting in the mPFC and mGluR5 agonists enhance NMDA receptor currents in vitro, suggesting that mGluR5 activation in IL may contribute to fear extinction. In the current study, rats injected with the mGluR5 antagonist 2-methyl-6-(phenylethyl)-pyridine (MPEP) systemically, or intra-IL, prior to extinction exhibited normal within-session extinction, but were impaired in their ability to recall extinction the following day. To directly determine whether mGluR5 stimulation enhances the burst firing of IL neurons, we used patch-clamp electrophysiology in prefrontal slices. The mGluR5 agonist, (RS)-2-chloro-5-hydroxyphenylglycine (CHPG), increased intrinsic bursting in IL neurons. Increased bursting was correlated with a reduction in the slow after hyperpolarizing potential and was prevented by coapplication of MPEP. CHPG did not increase NMDA currents, suggesting that an NMDA receptor-independent enhancement of IL bursting via stimulation of mGluR5 receptors contributes to fear extinction. Therefore, the mGluR5 receptor could be a suitable target for pharmacological adjuncts to extinction-based therapies for anxiety disorders.
The extinction of conditioned fear memories requires plasticity in the infralimbic medial prefrontal cortex (IL mPFC), but little is known about the molecular mechanisms involved. Brain-derived neurotrophic factor (BDNF) is a key mediator of synaptic plasticity in multiple brain areas. In rats subjected to auditory fear conditioning, BDNF infused into the IL mPFC reduced conditioned fear for up to 48 hours, even in the absence of extinction training, which suggests that BDNF substituted for extinction. Similar to extinction, BDNF-induced reduction in fear required N-methyl-D-aspartate receptors and did not erase the original fear memory. Rats failing to learn extinction showed reduced BDNF in hippocampal inputs to the IL mPFC, and augmenting BDNF in this pathway prevented extinction failure. Hence, boosting BDNF activity in hippocampal-infralimbic circuits may ameliorate disorders of learned fear.
Fear extinction is dependent on plasticity in the infralimbic prefrontal cortex, an area heavily innervated by midbrain dopaminergic inputs. Dopamine D2 receptors are concentrated in infralimbic output neurons that are involved in the suppression of conditioned fear after extinction. Here, we examined the specific role of infralimbic D2 receptors in mediating associative learning underlying fear extinction using the selective D2 antagonist raclopride.
Although fear research has largely focused on the amygdala, recent findings highlight cortical control of the amygdala in the service of fear regulation. In rodent models, it is becoming well established that the infralimbic (IL) prefrontal cortex plays a key role in extinction learning, and recent findings are uncovering molecular mechanisms involved in extinction-related plasticity. Furthermore, mounting evidence implicates the prelimbic (PL) prefrontal cortex in the production of fear responses. Both IL and PL integrate inputs from the amygdala, as well as other structures to gate the expression of fear via projections to inhibitory or excitatory circuits within the amygdala. We suggest that dual control of the amygdala by separate prefrontal modules increases the flexibility of an organisms response to danger cues.
There is current interest in identifying drugs that facilitate fear extinction, as this form of learning is the basis of certain cognitive therapies for anxiety disorders. Following an initial report several years ago that the alpha2-adrenoreceptor antagonist yohimbine facilitated extinction in mice, more recent studies have shown mixed effects or even impairment. It has become clear that the effect of yohimbine on extinction depends on a number of factors, including genetic background, contextual variables and the presence of competing behaviors. To what extent theses effects of yohimbine are mediated through the alpha2-adrenoreceptor, as opposed to other sites of action, is also uncertain. More work is needed before this drug can be approved as a pharmacological adjunct for extinction-based therapies. More generally, the case of yohimbine may serve as a model for the development of other extinction facilitators.
During auditory fear conditioning, it is well established that lateral amygdala (LA) neurons potentiate their response to the tone conditioned stimulus, and that this potentiation is required for conditioned fear behavior. Conditioned tone responses in LA, however, last only a few hundred milliseconds and cannot be responsible for sustained fear responses to a tone lasting tens of seconds. Recent evidence from inactivation and stimulation studies suggests that the prelimbic (PL) prefrontal cortex is necessary for expression of learned fears, but the timing of PL tone responses and correlations with fear behavior have not been studied. Using multichannel unit recording techniques in behaving rats, we observed sustained conditioned tone responses in PL that were correlated with freezing behavior on a second-to-second basis during the presentation of a 30 s tone. PL tone responses were also correlated with conditioned freezing across different experimental phases (habituation, conditioning, extinction). Moreover, the persistence of PL responses after extinction training was associated with failure to express extinction memory. Together with previous inactivation findings, the present results suggest that PL transforms transient amygdala inputs to a sustained output that drives conditioned fear responses and gates the expression of extinction. Given the relatively long latency of conditioned responses we observed in PL (approximately 100 ms after tone onset), we propose that PL integrates inputs from the amygdala, hippocampus, and other cortical sources to regulate the expression of fear memories.
The basolateral amygdala plays an important role in the acquisition and expression of both fear conditioning and fear extinction. To understand how a single structure could encode these "opposite" memories, we developed a biophysical network model of the lateral amygdala (LA) neurons during auditory fear conditioning and extinction. Membrane channel properties were selected to match waveforms and firing properties of pyramidal cells and interneurons in LA, from published in vitro studies. Hebbian plasticity was implemented in excitatory AMPA and inhibitory GABA(A) receptor-mediated synapses to model learning. The occurrence of synaptic potentiation versus depression was determined by intracellular calcium levels, according to the calcium control hypothesis. The model was able to replicate conditioning- and extinction-induced changes in tone responses of LA neurons in behaving rats. Our main finding is that LA activity during both acquisition and extinction can be controlled by a balance between pyramidal cell and interneuron activations. Extinction training depressed conditioned synapses and also potentiated local interneurons, thereby inhibiting the responses of pyramidal cells to auditory input. Both long-term depression and potentiation of inhibition were required to initiate and maintain extinction. The model provides insights into the sites of plasticity in conditioning and extinction, the mechanism of spontaneous recovery, and the role of amygdala NMDA receptors in extinction learning.
Although we know a great deal about how the brain processes information about aversive and appetitive stimuli, it is not clear how these two systems interact to guide behavior. In this issue of Neuron, Jhou and colleagues identify a region in the midbrain tegmentum that signals aversive events and inhibits midbrain dopamine neurons.
Psychostimulants, such as yohimbine and amphetamine, can enhance learning and memory. Extinction of conditioned fear involves new learning, so we asked whether psychostimulants could enhance this learning. Previous work suggests that yohimbine facilitates extinction, using freezing as a fear measure. However, psychostimulant-induced alterations in locomotion can confound freezing measurements. Furthermore, the effects of amphetamine on fear extinction have never been examined.
Previous work has implicated noradrenergic beta-receptors in the consolidation and reconsolidation of conditioned fear. Less is known, however, about their role in fear expression and extinction. The beta-receptor blocker propranolol has been used clinically to reduce anxiety. With an auditory fear conditioning task in rats, we assessed the effects of systemic propranolol on the expression and extinction of two measures of conditioned fear: freezing and suppression of bar-pressing.
Extinction is a form of inhibitory learning that suppresses a previously conditioned response. Both fear and drug seeking are conditioned responses that can lead to maladaptive behavior when expressed inappropriately, manifesting as anxiety disorders and addiction, respectively. Recent evidence indicates that the medial prefrontal cortex (mPFC) is critical for the extinction of both fear and drug-seeking behaviors. Moreover, a dorsal-ventral distinction is apparent within the mPFC, such that the prelimbic (PL-mPFC) cortex drives the expression of fear and drug seeking, whereas the infralimbic (IL-mPFC) cortex suppresses these behaviors after extinction. For conditioned fear, the dorsal-ventral dichotomy is accomplished via divergent projections to different subregions of the amygdala, whereas for drug seeking, it is accomplished via divergent projections to the subregions of the nucleus accumbens. Given that the mPFC represents a common node in the extinction circuit for these behaviors, treatments that target this region may help alleviate symptoms of both anxiety and addictive disorders by enhancing extinction memory.
The prefrontal cortex (PFC) regulates emotional responses, but it is unclear how PFC integrates diverse inputs to select the appropriate response. We therefore evaluated the contribution of basolateral amygdala (BLA) and ventral hippocampus (vHPC) inputs to fear signaling in the prelimbic (PL) cortex, a PFC region critical for the expression of conditioned fear. In conditioned rats trained to press for food, BLA inactivation decreased the activity of projection cells in PL, and reduced PL conditioned tone responses. In contrast, vHPC inactivation decreased activity of interneurons in PL and increased PL conditioned tone responses. Consistent with hippocampal gating of fear after extinction, vHPC inactivation increased fear and PL pyramidal activity in extinguished, but not in conditioned, rats. These results suggest a prefrontal circuit whereby hippocampus gates amygdala-based fear. Thus, deficient hippocampal inhibition of PFC may underlie emotional disorders, especially in light of reduced hippocampal volume observed in depression and PTSD.
Deep brain stimulation (DBS) of the ventral capsule/ventral striatum (VC/VS) reduces symptoms of intractable obsessive-compulsive disorder (OCD), but the mechanism of action is unknown. OCD is characterized by avoidance behaviors that fail to extinguish, and DBS could act, in part, by facilitating extinction of fear. We investigated this possibility by using auditory fear conditioning in rats, for which the circuits of fear extinction are well characterized. We found that DBS of the VS (the VC/VS homolog in rats) during extinction training reduced fear expression and strengthened extinction memory. Facilitation of extinction was observed for a specific zone of dorsomedial VS, just above the anterior commissure; stimulation of more ventrolateral sites in VS impaired extinction. DBS effects could not be obtained with pharmacological inactivation of either dorsomedial VS or ventrolateral VS, suggesting an extrastriatal mechanism. Accordingly, DBS of dorsomedial VS (but not ventrolateral VS) increased expression of a plasticity marker in the prelimbic and infralimbic prefrontal cortices, the orbitofrontal cortex, the amygdala central nucleus (lateral division), and intercalated cells, areas known to learn and express extinction. Facilitation of fear extinction suggests that, in accord with clinical observations, DBS could augment the effectiveness of cognitive behavioral therapies for OCD.
Individual differences in a persons ability to control fear have been linked to activation in the dorsal anterior cingulate cortex, the ventromedial prefrontal cortex, and the amygdala. This study investigated whether functional variance in this network can be predicted by resting metabolism in these same regions.
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