Classical Pavlovian Conditioning Research Articles

Higher threat avoidance costs reduce avoidance behaviour which in turn promotes fear extinction in humans

Theoretical models specifying the underlying mechanisms of the development and maintenance of anxiety and related disorders state that fear responses acquired through classical Pavlovian conditioning are maintained by repeated avoidance behaviour; thus, it is assumed that avoidance prevents fear extinction. The present study investigated behavioural avoidance decisions as a function of avoidance costs in a naturalistic fear conditioning paradigm. Ecologically valid avoidance costs - manipulated between participant groups - were represented via time-delays during a detour in a gamified computer task. After differential acquisitions of shock-expectancy to a predictive conditioned stimulus (CS+), participants underwent extinction where they could either take a risky shortcut, while anticipating shock signaled by the CS+, or choose a costly avoidance option (lengthy detour); thus, they were faced with an approach-avoidance conflict. Groups with higher avoidance costs (longer detours) showed lower proportions of avoiders. Avoiders gave heightened shock-expectancy ratings post-extinction, demonstrating ‘protecting from extinction’, i.e. failure to extinguish. Moreover, there was an indirect effect of avoidance costs on protection from extinction through avoidance behaviour. No moderating role of trait-anxiety was found. Theoretical implications of avoidance behaviour are discussed, considering the involvement of instrumental learning in the maintenance of fear responses.

Activity-dependent structural plasticity after aversive experiences in amygdala and auditory cortex pyramidal neurons

The brain is highly plastic and undergoes changes in response to many experiences. Learning especially can induce structural remodeling of dendritic spines, which is thought to relate to memory formation. Classical Pavlovian fear conditioning (FC) traditionally pairs an auditory cue with an aversive footshock, and has been widely used to study neural processes underlying associative learning and memory. Past research has found dendritic spine changes after FC in several structures. But, due to heterogeneity of cells within brain structures and limitations of traditional neuroanatomical techniques, it is unclear if all cells included in analyses were actually active during learning processes, even if known circuits are isolated. In this study, we employed a novel approach to analyze structural plasticity explicitly in neurons activated by exposure to either cued or uncued footshocks. We used male and female Arc-dVenus transgenic mice, which express the Venus fluorophore driven by the activity-related Arc promoter, to identify neurons that were active during either scenario. We then targeted fluorescent microinjections to Arc+ and neighboring Arc− neurons in the basolateral area of the amygdala (BLA) and auditory association cortex (TeA). In both BLA and TeA, Arc+ neurons had reduced thin and mushroom spine densities compared to Arc− neurons. This effect was present in males and females alike and also in both cued and uncued shock groups. Overall, this study adds to our understanding of how neuronal activity affects structural plasticity, and represents a methodological advance in the ways we can directly relate structural changes to experience-related neural activity.

Fight, Flight, or Freeze? The Answer May Depend on Your Sex

Pavlovian fear conditioning is one of the most common and well-understood methods for studying fear learning. However, research is predominantly performed in males. Recently, in a classical Pavlovian fear conditioning paradigm, Gruene and colleagues described an active conditioned fear response ('darting') prevalent in female rats that better maintain an extinction memory.

Abstract P201: Divergent Angiotensin Receptor Signaling in a Mouse Model of Post-Traumatic Stress Disorder (PTSD)

Independent of their beneficial effects on hypertension and cardiovascular related disease, angiotensin receptor type 1 (AT1R) blockers can improve stress-related symptoms. AT1R receptor-mediated actions can be counteracted directly or indirectly by the angiotensin receptor type 2 receptor (AT2R). Our recent studies in a mouse model of PTSD have shown that AT1R blockade increases the extinction (learned inhibition) of a traumatic fear memory and that AT1R mRNA expression is reduced in fear related brain regions of animals treated with the AT1R antagonist losartan. These data imply that downstream AT1 signaling events maybe important in consolidation of fear memory extinction. Therefore we investigated the acute effects of AT2R inhibition and AT2R stimulation on fear memory and baseline anxiety in mice. We performed classical Pavlovian fear conditioning pairing auditory cues with foot shocks and examined fear extinction (% freezing to conditioned stimulus) in the presence of the AT2R antagonist PD 123319 and agonist Compound 21. Twenty-four hours following fear conditioning, PD 123319 (15 mg/kg IP), Compound 21 (10 mg/kg IP) or vehicle was administered prior to fear memory extinction. The PD treated group exhibited significantly less percent freezing (68%; 68 of 100) compared to vehicle control (47%; 47 of 100) during fear expression (F10, 300 = 1.9; p<0.05, n=15) while no effect was observed during extinction retention, an index of long-term fear memory. On the other hand, Compound 21 had no effect on fear expression, extinction or basal levels of anxiety. Moreover, following fear conditioning, qPCR data revealed that mRNA expression of AT2R and angiotensin converting enzyme 2 (ACE2) in the central amygdala were elevated (5 fold and 3 fold respectively, P<0.05, n=6) compared to home cage control, however the AT1R and angiotensin converting enzyme (ACE) gene expression pathways were unaltered. PD 123319 and Compound 21 had no effect on basal levels of anxiety as determined by open field testing. These data indicate that AT1R and AT2R may have divergent effects on short and long-term fear memory formation. Further studies are required to understand the differential regulation of angiotensin receptor signaling in PTSD.

Neural signatures of human fear conditioning: an updated and extended meta-analysis of fMRI studies.

Classical Pavlovian fear conditioning remains the most widely employed experimental model of fear and anxiety, and continues to inform contemporary pathophysiological accounts of clinical anxiety disorders. Despite its widespread application in human and animal studies, the neurobiological basis of fear conditioning remains only partially understood. Here we provide a comprehensive meta-analysis of human fear-conditioning studies carried out with functional magnetic resonance imaging (fMRI), yielding a pooled sample of 677 participants from 27 independent studies. As a distinguishing feature of this meta-analysis, original statistical brain maps were obtained from the authors of 13 of these studies. Our primary analyses demonstrate that human fear conditioning is associated with a consistent and robust pattern of neural activation across a hypothesized genuine network of brain regions resembling existing anatomical descriptions of the 'central autonomic-interoceptive network'. This finding is discussed with a particular emphasis on the neural substrates of conscious fear processing. Our associated meta-analysis of functional deactivations-a scarcely addressed dynamic in fMRI fear-conditioning studies-also suggests the existence of a coordinated brain response potentially underlying the 'safety signal' (that is, non-threat) processing. We attempt to provide an integrated summary on these findings with the view that they may inform ongoing studies of fear-conditioning processes both in healthy and clinical populations, as investigated with neuroimaging and other experimental approaches.

Aberrant Angiotensin Signaling in a Mouse Model of Post‐Traumatic Stress Disorder (PTSD)

Independent of their beneficial effects on hypertension and cardiovascular related disease, angiotensin receptor type 1 (AT1R) blockers can improve stress-related symptoms. AT1R receptor-mediated actions can be counteracted directly or indirectly by the angiotensin receptor type 2 receptor (AT2R). Our recent studies in a mouse model of PTSD have shown that AT1R blockade increases the extinction (learned inhibition) of a traumatic fear memory and that AT1R mRNA expression is reduced in fear related brain regions of losartan treated animals. These data imply that downstream AT1 signaling events maybe important in consolidation of fear memory extinction. Therefore we investigated the acute effects of AT2R inhibition on fear memory and baseline anxiety. We performed classical Pavlovian fear conditioning pairing auditory cues with foot shocks and examined fear extinction behavior and cardiovascular responses in the presence of the AT2R antagonist PD 123319. Twenty-four hours following fear conditioning, PD 123319 (15 mg/kg IP) was administered prior to fear memory extinction. The PD treated group exhibited significantly less freezing behavior (F10, 300 = 1.9; p<0.05) during fear expression and contrary to our previous results with the AT1 antagonist losartan, there was no effect during extinction retention, an index of long-term fear memory. These data indicate that AT1R and AT2R may have divergent effects on short and long-term fear memory formation. Further studies are required to understand the differential regulation of angiotensin receptor signaling in PTSD.

The strength of aversive and appetitive associations and maladaptive behaviors.

Certain maladaptive behaviors are thought to be acquired through classical Pavlovian conditioning. Exaggerated fear response, which can develop through Pavlovian conditioning, is associated with acquired anxiety disorders such as post-traumatic stress disorders (PTSDs). Inflated reward-seeking behavior, which develops through Pavlovian conditioning, underlies some types of addictive behavior (e.g., addiction to drugs, food, and gambling). These maladaptive behaviors are dependent on associative learning and the development of long-term memory (LTM). In animal models, an aversive reinforcer (fear conditioning) encodes an aversive contextual and cued LTM. On the other hand, an appetitive reinforcer results in conditioned place preference (CPP) that encodes an appetitive contextual LTM. The literature on weak and strong associative learning pertaining to the development of aversive and appetitive LTM is relatively scarce; thus, this review is particularly focused on the strength of associative learning. The strength of associative learning is dependent on the valence of the reinforcer and the salience of the conditioned stimulus that ultimately sways the strength of the memory trace. Our studies suggest that labile (weak) aversive and appetitive LTM may share similar signaling pathways, whereas stable (strong) aversive and appetitive LTM is mediated through different pathways. In addition, we provide some evidence suggesting that extinction of aversive fear memory and appetitive drug memory is likely to be mediated through different signaling molecules. We put forward the importance of studies aimed to investigate the molecular mechanisms underlying the development of weak and strong memories (aversive and appetitive), which would ultimately help in the development of targeted pharmacotherapies for the management of maladaptive behaviors that arise from classical Pavlovian conditioning.

A potential role for the paraventricular nucleus of the thalamus in mediating individual variation in Pavlovian conditioned responses

There is ample evidence to suggest that the paraventricular nucleus of the thalamus (PVT) mediates cue-reward learning, especially as it relates to drug-seeking behavior. However, its exact role in these complex processes remains unknown. Here we will present and discuss data from our own laboratory which suggests that the PVT plays a role in multiple forms of stimulus-reward learning, and does so via distinct neurobiological systems. Using an animal model that captures individual variation in response to reward-associated cues, we are able to parse the incentive from the predictive properties of reward cues and to elucidate the neural circuitry underlying these different forms of cue-reward learning. When rats are exposed to a classical Pavlovian conditioning paradigm, wherein a cue predicts food reward, some rats, termed sign-trackers, approach and manipulate the cue upon its presentation. This behavior is indicative of attributing incentive salience to the cue. That is, the cue gains excessive control over behavior for sign-trackers. In contrast, other rats, termed goal-trackers, treat the cue as a mere predictor, and upon its presentation go to the location of reward delivery. Based on our own data utilizing this model, we hypothesize that the PVT represents a common node, but differentially regulates the sign- vs. goal-tracking response. We postulate that the PVT regulates sign-tracking behavior, or the attribution of incentive salience, via subcortical, dopamine-dependent mechanisms. In contrast, we propose that goal-tracking behavior, or the attribution of predictive value, is the product of “top-down” glutamatergic processing between the prelimbic cortex (PrL) and the PVT. Together, data from our laboratory and others support a role for the PVT in cue-motivated behaviors and suggest that it may be an important locus within the neural circuitry that goes awry in addiction and related disorders.

Programming a Pavlovian-like conditioning circuit in Escherichia coli

Synthetic genetic circuits are programmed in living cells to perform predetermined cellular functions. However, designing higher-order genetic circuits for sophisticated cellular activities remains a substantial challenge. Here we program a genetic circuit that executes Pavlovian-like conditioning, an archetypical sequential-logic function, in Escherichia coli. The circuit design is first specified by the subfunctions that are necessary for the single simultaneous conditioning, and is further genetically implemented using four function modules. During this process, quantitative analysis is applied to the optimization of the modules and fine-tuning of the interconnections. Analogous to classical Pavlovian conditioning, the resultant circuit enables the cells to respond to a certain stimulus only after a conditioning process. We show that, although the conditioning is digital in single cells, a dynamically progressive conditioning process emerges at the population level. This circuit, together with its rational design strategy, is a key step towards the implementation of more sophisticated cellular computing.

Galantamine reverses scopolamine-induced behavioral alterations in Dugesia tigrina

In planaria (Dugesia tigrina), scopolamine, a nonselective muscarinic receptor antagonist, induced distinct behaviors of attenuated motility and C-like hyperactivity. Planarian locomotor velocity (pLMV) displayed a dose-dependent negative correlation with scopolamine concentrations from 0.001 to 1.0 mM, and a further increase in scopolamine concentration to 2.25 mM did not further decrease pLMV. Planarian hyperactivity counts was dose-dependently increased following pretreatment with scopolamine concentrations from 0.001 to 0.5 mM and then decreased for scopolamine concentrations ≥ 1 mM. Planarian learning and memory investigated using classical Pavlovian conditioning experiments demonstrated that scopolamine (1 mM) negatively influenced associative learning indicated by a significant decrease in % positive behaviors from 86 % (control) to 14 % (1 mM scopolamine) and similarly altered memory retention, which is indicated by a decrease in % positive behaviors from 69 % (control) to 27 % (1 mM scopolamine). Galantamine demonstrated a complex behavior in planarian motility experiments since co-application of low concentrations of galantamine (0.001 and 0.01 mM) protected planaria against 1 mM scopolamine-induced motility impairments; however, pLMV was significantly decreased when planaria were tested in the presence of 0.1 mM galantamine alone. Effects of co-treatment of scopolamine and galantamine on memory retention in planaria via classical Pavlovian conditioning experiments showed that galantamine (0.01 mM) partially reversed scopolamine (1 mM)-induced memory deficits in planaria as the % positive behaviors increased from 27 to 63 %. The results demonstrate, for the first time in planaria, scopolamine's effects in causing learning and memory impairments and galantamine's ability in reversing scopolamine-induced memory impairments.

FROM THE NEUROBIOLOGY OF EXTINCTION TO IMPROVED CLINICAL TREATMENTS

The neural circuitry underlying the fear response is extremely well conserved across mammalian species, which has allowed for the rapid translation of research findings in rodent models of fear to therapeutic interventions in human populations. Many aspects of exposure-based psychotherapy treatments in humans, which are widely used in the treatment of PTSD, panic disorder, phobias, and other anxiety disorders, are closely paralleled by extinction training in rodent fear conditioning models. Here, we discuss how the neural circuitry of fear learning and extinction in rodent animal models may be used to understand the underlying neural circuitry of fear-related disorders, such as PTSD in humans. We examine the factors that contribute to the pathology and development of PTSD. Next, we will review how fear is measured in animal models using classical Pavlovian fear conditioning paradigms, as well as brain regions such as the amygdala, which are involved in the fear response across species. Finally, we highlight the following three systems involved in the extinction of fear, all of which represent promising avenues for therapeutic interventions in the clinic: (1) the role of the glutamatergic N-methyl-d-aspartate (NMDA) receptor, (2) the role of the brain-derived neurotrophic factor (BDNF)-tyrosine kinase B (TrkB) induced signaling pathway, and (3) the role of the renin-angiotensin system. The modulation of pathways underlying fear learning and extinction, such as the ones presented in this review, in combination with extinction-based exposure therapy, represents promising avenues for therapeutic intervention in the treatment of human fear related disorders.

Angiotensin Type 1 Receptor Inhibition Enhances the Extinction of Fear Memory

The current effective treatment options for posttraumatic stress disorder (PTSD) are limited, and therefore the need to explore new treatment strategies is critical. Pharmacological inhibition of the renin-angiotensin system is a common approach to treat hypertension, and emerging evidence highlights the importance of this pathway in stress and anxiety. A recent clinical study from our laboratory provides evidence supporting a role for the renin-angiotensin system in the regulation of the stress response in patients diagnosed with PTSD. With an animal model of PTSD and the selective angiotensin receptor type 1 (AT1) antagonist losartan, we investigated the acute and long-term effects of AT1 receptor inhibition on fear memory and baseline anxiety. After losartan treatment, we performed classical Pavlovian fear conditioning pairing auditory cues with footshocks and examined extinction behavior, gene expression changes in the brain, as well as neuroendocrine and cardiovascular responses. After cued fear conditioning, both acute and 2-week administration of losartan enhanced the consolidation of extinction memory but had no effect on fear acquisition, baseline anxiety, blood pressure, and neuroendocrine stress measures. Gene expression changes in the brain were also altered in mice treated with losartan for 2 weeks, in particular reduced amygdala AT1 receptor and bed nucleus of the stria terminalis c-Fos messenger RNA levels. These data suggest that AT1 receptor antagonism enhances the extinction of fear memory and therefore might be a beneficial therapy for PTSD patients who have impairments in extinction of aversive memories.

Differential roles of the dorsal and ventral hippocampus in predator odor contextual fear conditioning

The study of fear memory is important for understanding various anxiety disorders in which patients experience persistent recollections of traumatic events. These memories often involve associations of contextual cues with aversive events; consequently, Pavlovian classical conditioning is commonly used to study contextual fear learning. The use of predator odor as a fearful stimulus in contextual fear conditioning has become increasingly important as an animal model of anxiety disorders. Innate fear responses to predator odors are well characterized and reliable; however, attempts to use these odors as unconditioned stimuli in fear conditioning paradigms have proven inconsistent. Here we characterize a contextual fear conditioning paradigm using coyote urine as the unconditioned stimulus. We found that contextual conditioning induced by exposure to coyote urine produces long-term freezing, a stereotypic response to fear observed in mice. This paradigm is context-specific and parallels shock-induced contextual conditioning in that it is responsive to extinction training and manipulations of predator odor intensity. Region-specific lesions of the dorsal and ventral hippocampus indicate that both areas are independently required for the long-term expression of learned fear. These results in conjunction with c-fos immunostaining data suggest that while both the dorsal and ventral hippocampus are required for forming a contextual representation, the ventral region also modulates defensive behaviors associated with predators. This study provides information about the individual contributions of the dorsal and ventral hippocampus to ethologically relevant fear learning.

Mediation of the extinction process in behaviorally conditioned immunosuppression

To analyze neuro-immune-communication, our research group established a model of conditioned immunosuppression in rats which follows the principle of classical (Pavlovian) conditioning. We employed a conditioned taste aversion (CTA) paradigm where the novel taste saccharin (conditioned stimulus/CS) is paired with the immunosuppressive drug cyclosporine A (unconditioned stimulus/US). Upon re-exposure to the conditioned stimulus animals show a reduced CTA (saccharine intake), as well as a significant inhibition of T lymphocyte proliferation and IFN-γ and IL-2 in anti-CD3 stimulated splenic T cells. Repeated, unreinforced re-exposure to the CS leads to an extinction of the conditioned behavioral (CTA) and immune responses (cytokine inhibition, lymphocyte proliferation). In order to analyze the neurobiological mechanisms responsible for the extinction process we administered the protein synthesis inhibitor Anisomycin, or the beta-adrenergic receptor blocker Propranolol into the insular cortex during evocation, as the insular cortex is known to mediate the learned immune response during re-exposure of the CS. Our preliminary data show a prolonged extinction of the CTA reflected by a reduced CS intake after micro injection of protein synthesis inhibitor Anisomycin but not after administration of Propranolol into the insular cortex. Ongoing experiments will reveal whether the prolonged extinction on the behavioral level can also be detected in the conditioned immune response.

Fear Conditioning Assay in&amp;#160;Mouse

[Abstract] The study of fear memory is important for understanding various anxiety disorders in which patients experience persistent recollections of traumatic events. These memories often involve associations of contextual cues with aversive events; consequently, Pavlovian classical conditioning is commonly used to study contextual fear learning. A form of contextual fear conditioning that is becoming increasingly important as an animal model of anxiety disorders uses predator odor as a fearful stimulus. Innate fear responses to predator odors are well characterized and reliable; however, attempts to use these odors as unconditioned stimuli in fear conditioning paradigms have been highly dependent on experimental setup and have produced inconsistent behavioral results. Here we present a contextual fear conditioning paradigm using coyote urine as the unconditioned stimulus, which has been shown to produce consistent contextual freezing in response to fear learning (Wang et al., 2012).

Lessons from heartbreak

Male fruitflies quickly learn that courting already-mated females is useless. It turns out that a small subset of neurons in the male brain signals this negative experience and controls pheromone sensitivity. See Letter p.145 Courtship behaviour in Drosophila has become a leading model for defining the neural circuits and molecular mechanisms that underlie complex innate behaviours. Young male flies initially chase any female in the vicinity, but soon learn to avoid those perfumed with the male pheromone cis-vaccenyl acetate (cVA), which indicates loss of virginity. This study shows that this learning is not classical Pavlovian conditioning occurring through arbitrary association of cVA with rejection. Rather, it results from an increase in the males' innate sensitivity for cVA, and is controlled by a small circuit of dopaminergic neurons. The results pave the way for the study of the more sophisticated learning rules involved in sensory integration and fine decision-making.

Use of honey bees (Apis mellifera L.) to detect the presence of Mediterranean fruit fly (Ceratitis capitata Wiedemann) larvae in Valencia oranges

When fruit deteriorates a characteristic profile of volatile chemicals is produced that is different from that produced by healthy fruits. The identification of such chemicals allows the possibility of monitoring the fruit for early signs of deterioration with biological sensors. The use of honey bees and other insects as biological sensors is well known. This study aimed to identify the volatiles produced by oranges infested with larvae of the Mediterranean fruit fly and to test the ability of honey bees, conditioned to this volatile chemical profile, to detect such oranges. Seventeen compounds that were present in higher concentrations in the volatile profiles of infested oranges than in those of insect-free fruits were mixed at the same relative concentrations as those in the collected volatiles of infested oranges. The synthetic mixture was used to train honey bees by classical Pavlovian conditioning and subsequent tests showed that they were then able to discriminate between medfly-infested and uninfested oranges. This study demonstrates an innovative way of detecting, at an early stage, the symptoms of damage to oranges by the Mediterranean fruit fly.

What ethologically based models have taught us about the neural systems underlying fear and anxiety

Classical Pavlovian fear conditioning to painful stimuli has provided the generally accepted view of a core system centered in the central amygdala to organize fear responses. Ethologically based models using other sources of threat likely to be expected in a natural environment, such as predators or aggressive dominant conspecifics, have challenged this concept of a unitary core circuit for fear processing. We discuss here what the ethologically based models have told us about the neural systems organizing fear responses. We explored the concept that parallel paths process different classes of threats, and that these different paths influence distinct regions in the periaqueductal gray - a critical element for the organization of all kinds of fear responses. Despite this parallel processing of different kinds of threats, we have discussed an interesting emerging view that common cortical-hippocampal-amygdalar paths seem to be engaged in fear conditioning to painful stimuli, to predators and, perhaps, to aggressive dominant conspecifics as well. Overall, the aim of this review is to bring into focus a more global and comprehensive view of the systems organizing fear responses.

Adult-onset hypothyroidism enhances fear memory and upregulates mineralocorticoid and glucocorticoid receptors in the amygdala.

Hypothyroidism is the most common hormonal disease in adults, which is frequently accompanied by learning and memory impairments and emotional disorders. However, the deleterious effects of thyroid hormones deficiency on emotional memory are poorly understood and often underestimated. To evaluate the consequences of hypothyroidism on emotional learning and memory, we have performed a classical Pavlovian fear conditioning paradigm in euthyroid and adult-thyroidectomized Wistar rats. In this experimental model, learning acquisition was not impaired, fear memory was enhanced, memory extinction was delayed and spontaneous recovery of fear memory was exacerbated in hypothyroid rats. The potentiation of emotional memory under hypothyroidism was associated with an increase of corticosterone release after fear conditioning and with higher expression of glucocorticoid and mineralocorticoid receptors in the lateral and basolateral nuclei of the amygdala, nuclei that are critically involved in the circuitry of fear memory. Our results demonstrate for the first time that adult-onset hypothyroidism potentiates fear memory and also increases vulnerability to develop emotional memories. Furthermore, our findings suggest that enhanced corticosterone signaling in the amygdala is involved in the pathophysiological mechanisms of fear memory potentiation. Therefore, we recommend evaluating whether inappropriate regulation of fear in patients with post-traumatic stress and other mental disorders is associated with abnormal levels of thyroid hormones, especially those patients refractory to treatment.

A meta-analysis of instructed fear studies: Implications for conscious appraisal of threat

In classical Pavlovian fear conditioning, a neutral stimulus (conditioned stimulus, CS) comes to be evaluated as threatening due to its association with an aversive stimulus (unconditioned stimulus, UCS), and elicits fear. In a subtype of fear conditioning paradigms, called instructed fear or anticipatory anxiety, subjects are made aware of the CS–UCS association prior to actually experiencing it. Initial fear elicitation during this type of conditioning results from the negative evaluation of the CS as a consequence of CS–UCS contingency awareness. Prior reports have suggested that this conscious appraisal process is mediated by a variety of brain regions, including rostral dorsomedial prefrontal/dorsal anterior cingulate cortex (dmPFC/dACC), lateral prefrontal cortex (lPFC), posterior cingulate, hippocampus/parahippocampus, and nucleus accumbens, but there is little overlap between results. We reasoned that a formal meta-analysis of existing instructed fear studies should help narrow down the search for conscious appraisal areas in fear conditioning to those consistently activated across studies. We found consistent activation in rostral dmPFC but not in the other candidate areas. These results allow for maintaining the theory that the rostral dmPFC is involved in conscious threat appraisal. We also report a meta-analysis of uninstructed (classical) fear conditioning studies in which we found notable activation in more posterior parts of the dmPFC/dACC that overlapped with some of the instructed fear activations. These data suggest that mid regions of the dmPFC/dACC are part of a “core” fear network that is activated irrespective of how fear was learnt.