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Aversive learning can produce a wide variety of defensive behavioral responses depending on the circumstances, ranging from reactive responses like freezing to proactive avoidance responses. While most of this initial learning is behaviorally supported by an expectancy of an aversive outcome and neurally supported by activity within the basolateral amygdala, activity in other brain regions become necessary for the execution of defensive strategies that emerge in other aversive learning paradigms such as active avoidance. Here, we review the neural circuits that support both reactive and proactive defensive behaviors that are motivated by aversive learning, and identify commonalities between the neural substrates of these distinct (and often exclusive) behavioral strategies., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The ventral pallidum (VP) receives its primary inputs from the nucleus accumbens (NAC) and the basolateral amygdala (BLA). We demonstrated recently that in the VP, the D2 DA receptor (D 2 R) agonist quinpirole dose-dependently facilitates memory consolidation in inhibitory avoidance and spatial learning. In the VP, D 2 R can be found both on NAC and BLA terminals. According to our hypothesis, quinpirole microinjected into the VP can facilitate memory consolidation via modulation of synaptic plasticity on NAC and/or BLA terminals. The effect of intra-VP quinpirole on BLA-VP and NAC shell-VP synapses was investigated via a high frequency stimulation (HFS) protocol. Quinpirole was administered in three doses into the VP of male Sprague-Dawley rats after HFS; controls received vehicle. To examine whether an interaction between the NAC shell and the BLA at the level of the VP was involved, tetrodotoxin (TTX) was microinjected into one of the nuclei while stimulating the other nucleus. Our results showed that quinpirole dose-dependently modulates BLA-VP and NAC shell-VP synapses, similar to those observed in inhibitory avoidance and spatial learning, respectively. The lower dose inhibits BLA inputs, while the larger doses facilitates NAC shell inputs. The experiments with TTX demonstrates that the two nuclei do not influence each others' evoked responses in the VP. Power spectral density analysis demonstrated that independent from the synaptic facilitation, intra-VP quinpirole increases the amplitude of gamma frequency band after NAC HFS, and BLA tonically suppresses the NAC's HFS-induced gamma facilitation. In contrast, HFS of the BLA results in a delayed, transient increase in the amplitude of the gamma frequency band correlating with the LTP of the P1 component of the VP response to BLA stimulation. Furthermore, our results demonstrate that the BLA plays a prominent role in the generation of the delta oscillations: HFS of the BLA leads to a gradually increasing delta frequency band facilitation over time, while BLA inhibition blocks the NAC's HFS induced strong delta facilitation. These findings demonstrate that there is a complex interaction between the NAC shell region and the VP, as well as the BLA and the VP, and support the important role of VP D 2 Rs in the regulation of limbic information flow., Competing Interests: Declaration of competing interest Anthony A. Grace has received funds from Lundbeck, Pfizer, Lilly, Roche, Janssen, Alkermes, Newron, Takeda and Merck. The other author declares no conflict of interest., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

The sense of smell is tightly linked to emotions, a link that is thought to rely on the direct synaptic connections between the olfactory bulb (OB) and nuclei of the amygdala. However, there are multiple pathways projecting olfactory information to the amygdala, and their unique functions are unknown. The pathway via the nucleus of the lateral olfactory tract (NLOT) that receives input from olfactory regions and projects to the basolateral amygdala (BLA) is among them. NLOT has been very little studied, and consequentially its function is unknown. Furthermore, formulation of informed hypotheses about NLOT function is at this stage limited by the lack of knowledge about its connectivity and physiological properties. Here, we used virus-based tracing methods to systematically reveal inputs into NLOT, as well as NLOT projection targets in mice of both sexes. We found that the NLOT is interconnected with several olfactory brain regions and with the BLA. Some of these connections were reciprocal, and some showed unique interhemispheric patterns. We tested the excitable properties of NLOT neurons and the properties of each of the major synaptic inputs. We found that the NLOT receives powerful input from the piriform cortex, tenia tecta, and the BLA but only very weak input from the OB. When input crosses threshold, NLOT neurons respond with calcium-dependent bursts of action potentials. We hypothesize that this integration of olfactory and amygdalar inputs serves behaviors that combine smell and emotion., Competing Interests: The authors declare no competing financial interests., (Copyright © 2024 the authors.)

Pavlovian fear conditioning research suggests that the interaction between the dorsal periaqueductal gray (dPAG) and basolateral amygdala (BLA) acts as a prediction error mechanism in the formation of associative fear memories. However, their roles in responding to naturalistic predatory threats, characterized by less explicit cues and the absence of reiterative trial-and-error learning events, remain unexplored. In this study, we conducted single-unit recordings in rats during an 'approach food-avoid predator' task, focusing on the responsiveness of dPAG and BLA neurons to a rapidly approaching robot predator. Optogenetic stimulation of the dPAG triggered fleeing behaviors and increased BLA activity in naive rats. Notably, BLA neurons activated by dPAG stimulation displayed immediate responses to the robot, demonstrating heightened synchronous activity compared to BLA neurons that did not respond to dPAG stimulation. Additionally, the use of anterograde and retrograde tracer injections into the dPAG and BLA, respectively, coupled with c-Fos activation in response to predatory threats, indicates that the midline thalamus may play an intermediary role in innate antipredatory-defensive functioning., Competing Interests: EK, MK, SP, JC, JK No competing interests declared, (© 2023, Kim et al.)

The basolateral amygdala (BLA) contains discrete neuronal circuits that integrate positive or negative emotional information and drive the appropriate innate and learned behaviors. Whether these circuits consist of genetically-identifiable and anatomically segregated neuron types, is poorly understood. Also, our understanding of the response patterns and behavioral spectra of genetically-identifiable BLA neurons is limited. Here, we classified 11 glutamatergic cell clusters in mouse BLA and found that several of them were anatomically segregated in lateral versus basal amygdala, and anterior versus posterior regions of the BLA. Two of these BLA subpopulations innately responded to valence-specific, whereas one responded to mixed - aversive and social - cues. Positive-valence BLA neurons promoted normal feeding, while mixed selectivity neurons promoted fear learning and social interactions. These findings enhance our understanding of cell type diversity and spatial organization of the BLA and the role of distinct BLA populations in representing valence-specific and mixed stimuli., (© 2024. The Author(s).)

The basolateral amygdala (BLA) is the subregion of the amygdala located in the medial of the temporal lobe, which is connected with a wide range of brain regions to achieve diverse functions. Recently, an increasing number of studies have focused on the participation of the BLA in many neuropsychiatric disorders from the neural circuit perspective, aided by the rapid development of viral tracing methods and increasingly specific neural modulation technologies. However, how to translate this circuit-level preclinical intervention into clinical treatment using noninvasive or minor invasive manipulations to benefit patients struggling with neuropsychiatric disorders is still an inevitable question to be considered. In this review, we summarized the role of BLA-involved circuits in neuropsychiatric disorders including Alzheimer's disease, perioperative neurocognitive disorders, schizophrenia, anxiety disorders, depressive disorders, posttraumatic stress disorders, autism spectrum disorders, and pain-associative affective states and cognitive dysfunctions. Additionally, we provide insights into future directions and challenges for clinical translation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Objectives: Animal studies have indicated that fat intake mediates amygdala activation, which in turn promotes fat intake, while amygdala activation increases the preference for fat and leads to increased fat intake. However, the association among fat intake, amygdala activation, and appetite for high-calorie foods in humans remains unclear. Thus, to examine this association, we conducted a functional magnetic resonance imaging (fMRI) experiment., Methods: Fifty healthy-weight adults (18 females; mean age: 22.9 ± 3.02 years) were included. Participants were shown images of high-calorie and low-calorie foods and were instructed to rate their desire to eat the food items during fMRI. All participants provided information on their daily fat intake using a self-reported questionnaire. Associations among fat intake, the desire to eat high-calorie or low-calorie food items, and amygdala responses to food items were examined., Results: The basolateral amygdala (BLA) response was positively associated with fat intake ([ x , y , z ] = [24, -6, -16], z  = 3.91, p FWE-corrected  = 0.007) and the desire to eat high-calorie food items ([26, -4, -16], z  = 3.75, p FWE-corrected  = 0.010). Structural equation modeling showed that the desire for high-calorie food items was predicted by BLA response to high-calorie food items ( p  = 0.013, β  = 3.176), and BLA response was predicted by fat intake ( p  < 0.001, β  = 0.026)., Discussion: Fat intake influences BLA response to high-fat food, which in turn increases the desire to eat palatable high-fat food. This may lead to additional fat intake and increase the risk of weight gain.

It is well established that the basolateral amygdala (BLA) is an emotional processing hub that governs a diverse repertoire of behaviors. Selective engagement of a heterogeneous cell population in the BLA is thought to contribute to this flexibility in behavioral outcomes. However, whether this process is impacted by previous experiences that influence emotional processing remains unclear. Here we demonstrate that previous positive (enriched environment [EE]) or negative (chronic unpredictable stress [CUS]) experiences differentially influence the activity of populations of BLA principal neurons projecting to either the nucleus accumbens core or bed nucleus of the stria terminalis. Chemogenetic manipulation of these projection-specific neurons can mimic or occlude the effects of CUS and EE on behavioral outcomes to bidirectionally control avoidance behaviors and stress-induced helplessness. These data demonstrate that previous experiences influence the responsiveness of projection-specific BLA principal neurons, biasing information routing through the BLA, to drive divergent behavioral outcomes., Competing Interests: Declaration of interests J.L.M. serves as a member of the scientific advisory board for SAGE Therapeutics, Inc., for work unrelated to this project., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Basolateral amygdala (BLA) is a key hub for affect in the brain, 1 , 2 , 3 and dysfunction within this area contributes to a host of psychiatric disorders. 4 , 5 BLA is extensively and reciprocally interconnected with frontal cortex, 6 , 7 , 8 , 9 and some aspects of its function are evolutionarily conserved across rodents, anthropoid primates, and humans. 10 Neuron density in BLA is substantially lower in primates compared to murine rodents, 11 and frontal cortex (FC) is dramatically expanded in primates, particularly the more anterior granular and dysgranular areas. 12 , 13 , 14 Yet, how these anatomical differences influence the projection patterns of single BLA neurons to frontal cortex across rodents and primates is unknown. Using a barcoded connectomic approach, we assessed the single BLA neuron connections to frontal cortex in mice and macaques. We found that BLA neurons are more likely to project to multiple distinct parts of FC in mice than in macaques. Further, while single BLA neuron projections to nucleus accumbens were similarly organized in mice and macaques, BLA-FC connections differed substantially. Notably, BLA connections to subcallosal anterior cingulate cortex (scACC) in macaques were least likely to branch to other medial frontal cortex areas compared to perigenual ACC (pgACC). This pattern of connections was reversed in the mouse homologues of these areas, infralimbic and prelimbic cortex (IL and PL), mirroring functional differences between rodents and non-human primates. Taken together, these results indicate that BLA connections to FC are not linearly scaled from mice to macaques and instead the organization of single-neuron BLA connections is distinct between these species., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The Coronavirus disease (COVID-19) pandemic led to heightened anxiety in adolescents. The basolateral amygdala (BLA) and the nucleus accumbens (NAcc) are implicated in response to stress and may contribute to anxiety. The role of threat- and reward-related circuitry in adolescent anxiety during the COVID-19 pandemic, however, is not clear. Ninety-nine adolescents underwent resting-state fMRI ∼1 year before the pandemic. Following shelter-in-place orders, adolescents reported their perceived stress and, 1 month later, their anxiety. Generalized multivariate analyses identified BLA and NAcc seed-based whole-brain functional connectivity maps with perceived stress. In the resulting significant clusters, we examined the association between seed-based connectivityand subsequent anxiety. Perceived stress was associated with bilateral BLA and NAcc connectivity across distributed clusters that included prefrontal, limbic, temporal, and cerebellar regions. Several NAcc connectivity clusters located in ventromedial prefrontal, parahippocampal, and temporal cortices were positively associated with anxiety; NAcc connectivity with the inferior frontal gyrus was negatively associated. BLA connectivity was not associated with anxiety. These results underscore the integrative role of the NAcc in responding to acute stressors and its relation to anxiety in adolescents. Elucidating the involvement of subcortical-cortical circuitry in adolescents' capacity to respond adaptively to environmental challenges can inform treatment for anxiety-related disorders., (© The Author(s) 2024. Published by Oxford University Press.)

The lateral amygdala (LA) encodes fear memories by potentiating sensory inputs associated with threats and, in the process, recruits 10-30% of its neurons per fear memory engram. However, how the local network within the LA processes this information and whether it also plays a role in storing it are still largely unknown. Here, using ex vivo 12-patch-clamp and in vivo 32-electrode electrophysiological recordings in the LA of fear-conditioned rats, in combination with activity-dependent fluorescent and optogenetic tagging and recall, we identified a sparsely connected network between principal LA neurons that is organized in clusters. Fear conditioning specifically causes potentiation of synaptic connections between learning-recruited neurons. These findings of synaptic plasticity in an autoassociative excitatory network of the LA may suggest a basic principle through which a small number of pyramidal neurons could encode a large number of memories., (© 2024. The Author(s).)

The advent of general anesthesia (GA) has significant implications for clinical practice. However, the exact mechanisms underlying GA-induced transitions in consciousness remain elusive. Given some similarities between GA and sleep, the sleep-arousal neural nuclei and circuits involved in sleep-arousal, including the 5-HTergic system, could be implicated in GA. Herein, we utilized pharmacology, optogenetics, chemogenetics, fiber photometry, and retrograde tracing to demonstrate that both endogenous and exogenous activation of the 5-HTergic neural circuit between the dorsal raphe nucleus (DR) and basolateral amygdala (BLA) promotes arousal and facilitates recovery of consciousness from sevoflurane anesthesia. Notably, the 5-HT1A receptor within this pathway holds a pivotal role. Our findings will be conducive to substantially expanding our comprehension of the neural circuit mechanisms underlying sevoflurane anesthesia and provide a potential target for modulating consciousness, ultimately leading to a reduction in anesthetic dose requirements and side effects., Competing Interests: Declaration of Competing Interest This manuscript has not been submitted for publication elsewhere (other than BioRxiv), and the authors have no conflicts of interest to declare., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Growing evidence indicates a critical role of astrocytes in learning and memory. However, little is known about the role of basolateral amygdala complex (BLA-C) astrocytes in contextual fear conditioning (CFC), a paradigm relevant to understand and generate treatments for fear- and anxiety-related disorders. To get insights on the involvement of BLA-C astrocytes in fear memory, fluorocitrate (FLC), a reversible astroglial metabolic inhibitor, was applied at critical moments of the memory processing in order to target the acquisition, consolidation, retrieval and reconsolidation process of the fear memory. Adult Wistar male rats were bilaterally cannulated in BLA-C. Ten days later they were infused with different doses of FLC (0.5 or 1 nmol/0.5 µl) or saline before or after CFC and before or after retrieval. FLC impaired fear memory expression when administered before and shortly after CFC, but not one hour later. Infusion of FLC prior and after retrieval did not affect the memory. Our findings suggest that BLA-C astrocytes are critically involved in the acquisition/early consolidation of fear memory but not in the retrieval and reconsolidation. Furthermore, the extinction process was presumably not affected (considering that peri-retrieval administration could also affect this process)., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The pivotal role of the basolateral amygdala (BLA) in the emotional modulation of hippocampal plasticity and memory consolidation is well-established. Specifically, multiple studies have demonstrated that the activation of the noradrenergic (NA) system within the BLA governs these modulatory effects. However, most current evidence has been obtained by direct infusion of synthetic NA or beta-adrenergic agonists. In the present study, we aimed to investigate the effect of endogenous NA release in the BLA, induced by a natural aversive stimulus (coyote urine), on memory consolidation for a low-arousing, hippocampal-dependent task. Our experiments combined a weak object location task (OLT) version with subsequent mild predator odor exposure (POE). To investigate the role of endogenous NA in the BLA in memory modulation, a subset of the animals (Wistar rats) was treated with the non-selective beta-blocker propranolol at the end of the behavioral procedures. Hippocampal tissue was collected 90 min after drug infusion or after the OLT test, which was performed 24 h later. We used the obtained samples to estimate the levels of phosphorylated CREB (pCREB) and activity-regulated cytoskeleton-associated protein (Arc)-two molecular markers of experience-dependent changes in neuronal activity. The result suggests that POE has the potential to become a valuable behavioral paradigm for studying the interaction between BLA and the hippocampus in memory prioritization and selectivity.

Foraging decisions involve assessing potential risks and prioritizing food sources, which can be challenging when confronted with changing and conflicting circumstances. A crucial aspect of this decision-making process is the ability to actively overcome defensive reactions to threats and focus on achieving specific goals. The ventral pallidum (VP) and basolateral amygdala (BLA) are two brain regions that play key roles in regulating behavior motivated by either rewards or threats. However, it is unclear whether these regions are necessary in decision-making processes involving competing motivational drives during conflict. Our aim was to investigate the requirements of the VP and BLA for foraging choices in conflicts involving overcoming defensive responses. Here, we used a novel foraging task and pharmacological techniques to inactivate either the VP or BLA or to disconnect these brain regions before conducting a conflict test in male rats. Our findings showed that BLA is necessary for making risky choices during conflicts, whereas VP is necessary for invigorating the drive to obtain food, regardless of the presence of conflict. Importantly, our research revealed that the connection between VP and BLA is critical in controlling risky food-seeking choices during conflict situations. This study provides a new perspective on the collaborative function of VP and BLA in driving behavior, aimed at achieving goals in the face of dangers., Competing Interests: The authors declare no competing financial interests., (Copyright © 2024 the authors.)

The mammalian brain can store and retrieve memories of related events as distinct memories and remember common features of those experiences. How it computes this function remains elusive. Here, we show in rats that recent memories of two closely timed auditory fear events share overlapping neuronal ensembles in the basolateral amygdala (BLA) and are functionally linked. However, remote memories have reduced neuronal overlap and are functionally independent. The activity of parvalbumin (PV)-expressing neurons in the BLA plays a crucial role in forming separate remote memories. Chemogenetic blockade of PV preserves individual remote memories but prevents their segregation, resulting in reciprocal associations. The hippocampus drives this process through specific excitatory connections with BLA GABAergic interneurons. These findings provide insights into the neuronal mechanisms that minimize the overlap between distinct remote memories and enable the retrieval of related memories separately., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

The insular cortex, or insula, is a large brain region involved in the detection of thirst and the regulation of water intake. However, our understanding of the topographical, circuit, and molecular mechanisms for controlling water intake within the insula remains parcellated. We found that type-1 cannabinoid (CB 1 ) receptors in the insular cortex cells participate in the regulation of water intake and deconstructed the circuit mechanisms of this control. Topographically, we revealed that the activity of excitatory neurons in both the anterior insula (aIC) and posterior insula (pIC) increases in response to water intake, yet only the specific removal of CB 1 receptors in the pIC decreases water intake. Interestingly, we found that CB 1 receptors are highly expressed in insula projections to the basolateral amygdala (BLA), while undetectable in the neighboring central part of the amygdala. Thus, we recorded the neurons of the aIC or pIC targeting the BLA (aIC-BLA and pIC-BLA) and found that they decreased their activity upon water drinking. Additionally, chemogenetic activation of pIC-BLA projection neurons decreased water intake. Finally, we uncovered CB 1 -dependent short-term synaptic plasticity (depolarization-induced suppression of excitation [DSE]) selectively in pIC-BLA, compared with aIC-BLA synapses. Altogether, our results support a model where CB 1 receptor signaling promotes water intake by inhibiting the pIC-BLA pathway, thereby contributing to the fine top-down control of thirst responses., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Memories are stored in engram cells, which are necessary and sufficient for memory recall. Recalling a memory might undergo reconsolidation or extinction. It has been suggested that the original memory engram is reactivated during reconsolidation so that memory can be updated. Conversely, during extinction training, a new memory is formed that suppresses the original engram. Nonetheless, it is unknown whether extinction creates a new engram or modifies the original fear engram. In this study, we utilized the Daun02 procedure, which uses c-Fos-lacZ rats to induce apoptosis of strongly activated neurons and examine whether a new memory trace emerges as a result of a short or long reactivation, or if these processes rely on modifications within the original engram located in the basolateral amygdala (BLA) and infralimbic (IL) cortex. By eliminating neurons activated during consolidation and reactivation, we observed significant impacts on fear memory, highlighting the importance of the BLA engram in these processes. Although we were unable to show any impact when removing the neurons activated after the test of a previously extinguished memory in the BLA, disrupting the IL extinction engram reactivated the aversive memory that was suppressed by the extinction memory. Thus, we demonstrated that the IL cortex plays a crucial role in the network involved in extinction, and disrupting this specific node alone is sufficient to impair extinction behavior. Additionally, our findings indicate that extinction memories rely on the formation of a new memory, supporting the theory that extinction memories rely on the formation of a new memory, whereas the reconsolidation process reactivates the same original memory trace., (© 2024 Wiley Periodicals LLC.)

Acetylcholine (ACh) is released from basal forebrain cholinergic neurons in response to salient stimuli and engages brain states supporting attention and memory. These high ACh states are associated with theta oscillations, which synchronize neuronal ensembles. Theta oscillations in the basolateral amygdala (BLA) in both humans and rodents have been shown to underlie emotional memory, yet their mechanism remains unclear. Here, using brain slice electrophysiology in male and female mice, we show large ACh stimuli evoke prolonged theta oscillations in BLA local field potentials that depend upon M3 muscarinic receptor activation of cholecystokinin (CCK) interneurons (INs) without the need for external glutamate signaling. Somatostatin (SOM) INs inhibit CCK INs and are themselves inhibited by ACh, providing a functional SOM→CCK IN circuit connection gating BLA theta. Parvalbumin (PV) INs, which can drive BLA oscillations in baseline states, are not involved in the generation of ACh-induced theta, highlighting that ACh induces a cellular switch in the control of BLA oscillatory activity and establishes an internally BLA-driven theta oscillation through CCK INs. Theta activity is more readily evoked in BLA over the cortex or hippocampus, suggesting preferential activation of the BLA during high ACh states. These data reveal a SOM→CCK IN circuit in the BLA that gates internal theta oscillations and suggest a mechanism by which salient stimuli acting through ACh switch the BLA into a network state enabling emotional memory., Competing Interests: The authors declare no competing financial interests., (Copyright © 2024 the authors.)

Understanding how distinct functional circuits are coordinated to fine-tune mood and behavior is of fundamental importance. Here, we observe that within the dense projections from basolateral amygdala (BLA) to bed nucleus of stria terminalis (BNST), there are two functionally opposing pathways orchestrated to enable contextually appropriate expression of anxiety-like behaviors in male mice. Specifically, the anterior BLA neurons predominantly innervate the anterodorsal BNST (adBNST), while their posterior counterparts send massive fibers to oval BNST (ovBNST) with moderate to adBNST. Optogenetic activation of the anterior and posterior BLA inputs oppositely regulated the activity of adBNST neurons and anxiety-like behaviors, via disengaging and engaging the inhibitory ovBNST-to-adBNST microcircuit, respectively. Importantly, the two pathways exhibited synchronized but opposite responses to both anxiolytic and anxiogenic stimuli, partially due to their mutual inhibition within BLA and the different inputs they receive. These findings reveal synergistic interactions between two BLA-to-BNST pathways for appropriate anxiety expression with ongoing environmental demands., (© 2024. The Author(s).)

Social experiences carry tremendous weight in our decision-making, even when social partners are not present. To determine mechanisms, we trained female mice to respond for two food reinforcers. Then, one food was paired with a novel conspecific. Mice later favored the conspecific-associated food, even in the absence of the conspecific. Chemogenetically silencing projections from the prelimbic subregion (PL) of the medial prefrontal cortex to the basolateral amygdala (BLA) obstructed this preference while leaving social discrimination intact, indicating that these projections are necessary for socially driven choice. Further, mice that performed the task had greater densities of dendritic spines on excitatory BLA neurons relative to mice that did not. We next induced chemogenetic receptors in cells active during social interactions-when mice were encoding information that impacted later behavior. BLA neurons stimulated by social experience were necessary for mice to later favor rewards associated with social conspecifics but not make other choices. This profile contrasted with that of PL neurons stimulated by social experience, which were necessary for choice behavior in social and nonsocial contexts alike. The PL may convey a generalized signal allowing mice to favor particular rewards, while units in the BLA process more specialized information, together supporting choice motivated by social information., (Copyright © 2024 the authors.)

The basolateral amygdala (BLA) mediates both fear and reward learning. 1 , 2 Previous work has shown that parvalbumin (PV) interneurons in the BLA contribute to BLA oscillatory states integral to fear expression. 3 , 4 , 5 , 6 , 7 However, despite it being critical to our understanding of reward behaviors, it is unknown whether BLA oscillatory states and PV interneurons similarly contribute to reward processing. Local field potentials in the BLA were collected as male and female mice consumed sucrose reward, where prominent changes in the beta band (15-30 Hz) emerged with reward experience. During consumption of one water bottle during a two-water-bottle choice test, rhythmic optogenetic stimulation of BLA PVs produced a robust bottle preference, showing that PVs can sufficiently drive reward seeking. Finally, to demonstrate that PV activity is necessary for reward value use, PVs were chemogenetically inhibited following outcome devaluation, rendering mice incapable of using updated reward representations to guide their behavior. Taken together, these experiments provide novel information about the physiological signatures of reward while highlighting BLA PV interneuron contributions to behaviors that are BLA dependent. This work builds upon established knowledge of PV involvement in fear expression and provides evidence that PV orchestration of unique BLA network states is involved in both learning types., Competing Interests: Declaration of interests J.L.M. serves as a member of the scientific advisory board for SAGE Therapeutics, Inc. for work unrelated to this project., (Copyright © 2024 Elsevier Inc. All rights reserved.)

To make adaptive decisions, we build an internal model of the associative relationships in an environment and use it to make predictions and inferences about specific available outcomes. Detailed, identity-specific cue-reward memories are a core feature of such cognitive maps. Here we used fiber photometry, cell-type and pathway-specific optogenetic manipulation, Pavlovian cue-reward conditioning and decision-making tests in male and female rats, to reveal that ventral tegmental area dopamine (VTA DA ) projections to the basolateral amygdala (BLA) drive the encoding of identity-specific cue-reward memories. Dopamine is released in the BLA during cue-reward pairing; VTA DA →BLA activity is necessary and sufficient to link the identifying features of a reward to a predictive cue but does not assign general incentive properties to the cue or mediate reinforcement. These data reveal a dopaminergic pathway for the learning that supports adaptive decision-making and help explain how VTA DA neurons achieve their emerging multifaceted role in learning., (© 2024. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Obesity results from excessive caloric input associated with overeating and presents a major public health challenge. The hypothalamus has received significant attention for its role in governing feeding behavior and body weight homeostasis. However, extrahypothalamic brain circuits also regulate appetite and consumption by altering sensory perception, motivation, and reward. We recently discovered a population of basal forebrain cholinergic (BFc) neurons that regulate appetite suppression. Through viral tracing methods in the mouse model, we found that BFc neurons densely innervate the basolateral amygdala (BLA), a limbic structure involved in motivated behaviors. Using channelrhodopsin-assisted circuit mapping, we identified cholinergic responses in BLA neurons following BFc circuit manipulations. Furthermore, in vivo acetylcholine sensor and genetically encoded calcium indicator imaging within the BLA (using GACh3 and GCaMP, respectively) revealed selective response patterns of activity during feeding. Finally, through optogenetic manipulations in vivo, we found that increased cholinergic signaling from the BFc to the BLA suppresses appetite and food intake. Together, these data support a model in which cholinergic signaling from the BFc to the BLA directly influences appetite and feeding behavior., Competing Interests: The authors declare no competing financial interests., (Copyright © 2024 Ortiz-Guzman et al.)

Memory encodes past experiences, thereby enabling future plans. The basolateral amygdala is a centre of salience networks that underlie emotional experiences and thus has a key role in long-term fear memory formation 1 . Here we used spatial and single-cell transcriptomics to illuminate the cellular and molecular architecture of the role of the basolateral amygdala in long-term memory. We identified transcriptional signatures in subpopulations of neurons and astrocytes that were memory-specific and persisted for weeks. These transcriptional signatures implicate neuropeptide and BDNF signalling, MAPK and CREB activation, ubiquitination pathways, and synaptic connectivity as key components of long-term memory. Notably, upon long-term memory formation, a neuronal subpopulation defined by increased Penk and decreased Tac expression constituted the most prominent component of the memory engram of the basolateral amygdala. These transcriptional changes were observed both with single-cell RNA sequencing and with single-molecule spatial transcriptomics in intact slices, thereby providing a rich spatial map of a memory engram. The spatial data enabled us to determine that this neuronal subpopulation interacts with adjacent astrocytes, and functional experiments show that neurons require interactions with astrocytes to encode long-term memory., (© 2024. The Author(s).)

It is widely accepted that fear memories are consolidated through protein synthesis-dependent changes in the basolateral amygdala complex (BLA). However, recent studies show that protein synthesis is not required to consolidate the memory of a new dangerous experience when it is similar to a prior experience. Here, we examined whether the protein synthesis requirement for consolidating the new experience varies with its spatial and temporal distance from the prior experience. Female and male rats were conditioned to fear a stimulus (S1, e.g., light) paired with shock in stage 1 and a second stimulus (S2, e.g., tone) that preceded additional S1-shock pairings (S2-S1-shock) in stage 2. The latter stage was followed by a BLA infusion of a protein synthesis inhibitor, cycloheximide, or vehicle. Subsequent testing with S2 revealed that protein synthesis in the BLA was not required to consolidate fear to S2 when the training stages occurred 48 h apart in the same context; was required when they were separated by 14 d or occurred in different contexts; but was again not required if S1 was re-presented after the delay or in the different context. Similarly, protein synthesis in the BLA was not required to re consolidate fear to S2 when the training stages occurred 48 h apart but was required when they occurred 14 d apart. Thus, the protein synthesis requirement for consolidating/reconsolidating fear memories in the BLA is determined by similarity between present and past experiences, the time and place in which they occur, and reminders of the past experiences., Competing Interests: The authors declare no competing financial interests., (Copyright © 2024 the authors.)

In this issue of Neuron, Piantadosi et al. 1 demonstrate that by precisely controlling the activity of individual negative-valence neurons and positive-valence neurons in the basolateral amygdala, one can alter animals' appetitive or aversive responses, respectively, establishing a causal role of these neurons in valence-specific behavior., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The basolateral amygdala (BLA) is an evolutionarily conserved brain region, well known for valence processing. Despite this central role, the relationship between activity of BLA neuronal ensembles in response to appetitive and aversive stimuli and the subsequent expression of valence-specific behavior has remained elusive. Here, we leverage two-photon calcium imaging combined with single-cell holographic photostimulation through an endoscopic lens to demonstrate a direct causal role for opposing ensembles of BLA neurons in the control of oppositely valenced behavior in mice. We report that targeted photostimulation of either appetitive or aversive BLA ensembles results in mutual inhibition and shifts behavioral responses to promote consumption of an aversive tastant or reduce consumption of an appetitive tastant, respectively. Here, we identify that neuronal encoding of valence in the BLA is graded and relies on the relative proportion of individual BLA neurons recruited in a stable appetitive or quinine ensemble., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Despite recent advancements in identifying engram cells, our understanding of their regulatory and functional mechanisms remains in its infancy. To provide mechanistic insight into engram cell functioning, we introduced a novel local microcircuit labeling technique that enables the labeling of intraregional synaptic connections. Utilizing this approach, we discovered a unique population of somatostatin (SOM) interneurons in the mouse basolateral amygdala (BLA). These neurons are activated during fear memory formation and exhibit a preference for forming synapses with excitatory engram neurons. Post-activation, these SOM neurons displayed varying excitability based on fear memory retrieval. Furthermore, when we modulated these SOM neurons chemogenetically, we observed changes in the expression of fear-related behaviors, both in a fear-associated context and in a novel setting. Our findings suggest that these activated SOM interneurons play a pivotal role in modulating engram cell activity. They influence the expression of fear-related behaviors through a mechanism that is dependent on memory cues., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

The amygdala is known to mediate in moderating the impacts of emotional arousal and stress on memory. According to a growing body of research, the basolateral amygdala (BLA) is an important locus for integrating neuromodulator influences coordinating the retrieval of different types of memory and anxiety. This study aimed to investigate how the epinephrine in the BLA affects hippocampal fear memory, anxiety, and plasticity in control and stressed rats. For four days, male Wistar rats were exposed to electrical foot-shock stress. Animals received bilateral micro-injections of either vehicle or epinephrine (1 µg/side) into the BLA over four days (5 min before foot-shock stress). Behavioral characteristics (fear memory and anxiety-like behavior), histological features and electrophysiological parameters were investigated. Epinephrine injection into BLA resulted in a considerable impairment of fear memory in stressed rats. On the other hand, epinephrine effectively affected fear memory in control rats. Under stress conditions, epinephrine in the BLA is thought to increase anxiety-like behaviors. Treatment with epinephrine significantly increases the slope of fEPSP in the CA1 region of the hippocampus in the control and stress rats. In different groups, foot-shock stress had no effect on the apical and basal dendritic length in the CA1 region of the hippocampus. These results indicate that activating adrenergic receptors diminish fear memory and anxiety-like behaviors in the foot-shock stress, which this impact is independent of CA1 long-term potentiation induction., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Activity in the basolateral amygdala complex (BLA) is needed to encode fears acquired through contact with both innate sources of danger (i.e., things that are painful) and learned sources of danger (e.g., being threatened with a gun). However, within the BLA, the molecular processes required to consolidate the two types of fear are not the same: protein synthesis is needed to consolidate the first type of fear (so-called first-order fear) but not the latter (so-called second-order fear). The present study examined why first- and second-order fears differ in this respect. Specifically, it used a range of conditioning protocols in male and female rats, and assessed the effects of a BLA infusion of the protein synthesis inhibitor, cycloheximide, on first- and second-order conditioned fear. The results revealed that the differential protein synthesis requirements for consolidation of first- and second-order fears reflect differences in what is learned in each case. Protein synthesis in the BLA is needed to consolidate fears that result from encoding of relations between stimuli in the environment (stimulus-stimulus associations, typical for first-order fear) but is not needed to consolidate fears that form when environmental stimuli associate directly with fear responses emitted by the animal (stimulus-response associations, typical for second-order fear). Thus, the substrates of Pavlovian fear conditioning in the BLA depend on the way that the environment impinges upon the animal. This is discussed with respect to theories of amygdala function in Pavlovian fear conditioning, and ways in which stimulus-response associations might be consolidated in the brain., (Copyright © 2024 the authors.)

Reversal learning measures the ability to form flexible associations between choice outcomes with stimuli and actions that precede them. This type of learning is thought to rely on several cortical and subcortical areas, including the highly interconnected orbitofrontal cortex (OFC) and basolateral amygdala (BLA), and is often impaired in various neuropsychiatric and substance use disorders. However, the unique contributions of these regions to stimulus- and action-based reversal learning have not been systematically compared using a chemogenetic approach particularly before and after the first reversal that introduces new uncertainty. Here, we examined the roles of ventrolateral OFC (vlOFC) and BLA during reversal learning. Male and female rats were prepared with inhibitory designer receptors exclusively activated by designer drugs targeting projection neurons in these regions and tested on a series of deterministic and probabilistic reversals during which they learned about stimulus identity or side (left or right) associated with different reward probabilities. Using a counterbalanced within-subject design, we inhibited these regions prior to reversal sessions. We assessed initial and pre-/post-reversal changes in performance to measure learning and adjustments to reversals, respectively. We found that inhibition of the ventrolateral orbitofrontal cortex (vlOFC), but not BLA, eliminated adjustments to stimulus-based reversals. Inhibition of BLA, but not vlOFC, selectively impaired action-based probabilistic reversal learning, leaving deterministic reversal learning intact. vlOFC exhibited a sex-dependent role in early adjustment to action-based reversals, but not in overall learning. These results reveal dissociable roles for BLA and vlOFC in flexible learning and highlight a more crucial role for BLA in learning meaningful changes in the reward environment., (Copyright © 2024 the authors.)

The basolateral amygdaloid complex (BLA) is critically involved in emotional behaviors, such as aversive memory formation. In particular, fear memory after cued fear conditioning is strongly associated with the BLA, whereas both the BLA and hippocampus are essential for contextual fear memory formation. In the present study, we examined the effects of acute (3 h) sleep deprivation (SD) on BLA-associated fear memory in juvenile (P24-32) rats and performed in vitro electrophysiology using whole-cell patch clamping from the basolateral nucleus (BA) of the BLA. BA projection neurons exhibit the network oscillation, i.e., spontaneous oscillatory bursts of inhibitory transmission at 0.1-3 Hz, as previously reported. In the present study, SD either before or after fear conditioning (FC) disturbed the acquisition of tone-associated fear memory without significant effects on contextual fear memory. FC reduced the power of the oscillatory activity, but SD did not further reduce the oscillation power. Oscillation power was correlated with tone-associated freezing rate (FR) in SD-free fear-conditioned rats, but this relation was disrupted in SD treated group. Rhythm index (RI), the rhythmicity of the oscillation, quantified by autocorrelation analysis, also correlated with tone-associated FR in the combined data, including FC alone and FC with SD. These results suggest that slow network oscillation in the amygdala contributes to the formation of amygdala-dependent fear memory in relation to sleep., (Copyright © 2023 IBRO. Published by Elsevier Ltd. All rights reserved.)

Introduction: Relapse is a major treatment barrier for opioid use disorder. Environmental cues become associated with the rewarding effects of opioids and can precipitate relapse, even after numerous unreinforced cue presentations, due to deficits in extinction memory recall (EMR). Estradiol (E2) modulates EMR of fear-related cues, but it is unknown whether E2 impacts EMR of reward cues and what brain region(s) are responsible for E2s effects. Here, we hypothesize that inhibition of E2 signaling in the basolateral amygdala (BLA) will impair EMR of a heroin-associated cue in both male and female rats., Methods: We pharmacologically manipulated E2 signaling to characterize the role of E2 in the BLA on heroin-cue EMR. Following heroin self-administration, during which a light/tone cue was co-presented with each heroin infusion, rats underwent cued extinction to extinguish the conditioned association between the light/tone and heroin. During extinction, E2 signaling in the BLA was blocked by an aromatase inhibitor or specific estrogen receptor (ER) antagonists. The next day, subjects underwent a cued test to assess heroin-cue EMR., Results: In both experiments, females took more heroin than males (mg/kg) and had higher operant responding during cued extinction. Inhibition of E2 synthesis in the BLA impaired heroin-cue EMR in both sexes. Notably, E2s actions are mediated by different ER mechanisms, ERα in males but ERβ in females., Conclusions: This study is the first to demonstrate a behavioral role for centrally-produced E2 in the BLA and that E2 also impacts EMR of reward-associated stimuli in both sexes., (© 2023 S. Karger AG, Basel.)

Aims: The basolateral amygdala (BLA) plays an integral role in anxiety disorders (such as post traumatic stress disorder) stem from dysregulated fear memory. The excitability of glutamatergic neurons in the BLA correlates with fear memory, and the afterhyperpolarization current (I AHP ) mediated by small-conductance calcium-activated potassium channel subtype 2 (SK2) dominates the excitability of glutamatergicneurons. This study aimed to explore the effect of MPP2 interacts with SK2 in the excitability of glutamatergic neurons in the BLA and the extinction of conditioned fear in mice., Methods: Fear memory was analyzed via freezing percentage. Western blotting and fluorescence quantitative PCR were used to determine the expression of protein and mRNA respectively. Electrophysiology was employed to measure the excitability of glutamatergic neurons and I AHP ., Results: Fear conditioning decreased the levels of synaptic SK2 channels in the BLA, which were restored following fear extinction. Notably, reduced expression of synaptic SK2 channels in the BLA during fear conditioning was caused by the increased activity of protein kinase A (PKA), while increased levels of synaptic SK2 channels in the BLA during fear extinction were mediated by interactions with membrane-palmitoylated protein 2 (MPP2)., Conclusions: Our results revealed that MPP2 interacts with the SK2 channels and rescues the excitability of glutamatergic neurons by increasing the expression of synaptic SK2 channels in the BLA to promote the normalization of anxiety disorders and provide a new direction for the treatment., (© 2023 The Authors. CNS Neuroscience & Therapeutics Published by John Wiley & Sons Ltd.)

In trace fear conditioning, the prelimbic cortex exhibits persistent activity during the interval between the conditioned and unconditioned stimuli, which maintains a conditioned stimulus representation. Regions cooperating for this function or encoding the conditioned stimulus before the interval could send inputs to the prelimbic cortex, supporting learning. The basolateral amygdala has conditioned stimulus- and unconditioned stimulus-responsive neurons, convergently activated. The prelimbic cortex could directly project to the basolateral amygdala to associate the transient memory of the conditioned stimulus with the unconditioned stimulus. We investigated the neuronal circuit supporting temporal associations using contextual fear conditioning with a 5-s interval, in which 5 s separates the contextual conditioned stimulus from the unconditioned stimulus. Injecting retrobeads, we quantified c-Fos in prelimbic cortex- or basolateral amygdala-projecting neurons from 9 regions after contextual fear conditioning with a 5-s interval or contextual fear conditioning, in which the conditioned and unconditioned stimuli overlap. The contextual fear conditioning with a 5-s interval activated ventral CA1 and perirhinal cortex neurons projecting to the prelimbic cortex and prelimbic cortex neurons projecting to basolateral amygdala. Both fear conditioning activated ventral CA1 and lateral entorhinal cortex neurons projecting to basolateral amygdala and basolateral amygdala neurons projecting to prelimbic cortex. The perirhinal cortex → prelimbic cortex and ventral CA1 → prelimbic cortex connections are the first identified prelimbic cortex afferent projections participating in temporal associations. These results help to understand time-linked memories, a process required in episodic and working memories., (© The Author(s) 2023. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permission@oup.com.)

Reward devaluation adaptively controls reward intake. It remains unclear how cortical circuits causally encode reward devaluation in healthy and depressed states. Here, we show that the neural pathway from the anterior cingulate cortex (ACC) to the basolateral amygdala (BLA) employs a dynamic inhibition code to control reward devaluation and depression. Fiber photometry and imaging of ACC pyramidal neurons reveal reward-induced inhibition, which weakens during satiation and becomes further attenuated in depression mouse models. Ablating or inhibiting these neurons desensitizes reward devaluation, causes reward intake increase and ultimate obesity, and ameliorates depression, whereas activating the cells sensitizes reward devaluation, suppresses reward consumption, and produces depression-like behaviors. Among various ACC neuron subpopulations, the BLA-projecting subset bidirectionally regulates reward devaluation and depression-like behaviors. Our study thus uncovers a corticoamygdalar circuit that encodes reward devaluation via blunted inhibition and suggests that enhancing inhibition within this circuit may offer a therapeutic approach for treating depression., Competing Interests: Declaration of interests M.L. serves on Neuron’s scientific advisory board. M.L., Z.Y., and Z.Q. filed a patent related to this work., (Copyright © 2023 Elsevier Inc. All rights reserved.)

In female rats and humans, reproductive experience (i.e., pregnancy) alters the behavioral, hormonal and molecular substrates of fear extinction. Here, we assessed whether the role of a central neural substrate of fear extinction, the basolateral amygdala (BLA), also changes following reproductive experience. Nulliparous (virgin) and primiparous (one prior pregnancy) female rats received infusions of the GABA A agonist, muscimol, to temporarily inactivate the BLA prior to fear conditioning or extinction training. In follow up experiments, the BLA was also inactivated immediately after extinction training. BLA inactivation impaired the acquisition and expression of conditioned fear in both nulliparous and primiparous rats. In nulliparous rats, BLA inactivation prior to or immediately after extinction training impaired extinction retention. In contrast, in primiparous rats, BLA inactivation prior to or immediately after extinction training did not impair extinction retention, despite suppressing freezing during extinction training. These results suggest that, consistent with past findings in males, the BLA is a central component of the neural circuitry of fear acquisition and its extinction in virgin female rats. However, after pregnancy, female rats no longer depend on the BLA to extinguish fear, despite requiring the BLA to acquire conditioned fear. Given that fear extinction forms the basis of exposure therapy for anxiety disorders in humans, the present findings may have clinical implications. To improve the efficacy of exposure therapy for anxiety disorders, we may need to target different mechanisms in females dependent on their reproductive history., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Psychological loss is a common experience that erodes well-being and negatively impacts quality of life. The molecular underpinnings of loss are poorly understood. Here, we investigate the mechanisms of loss using an environmental enrichment removal (ER) paradigm in male rats. The basolateral amygdala (BLA) was identified as a region of interest, demonstrating differential Fos responsivity to ER and having an established role in stress processing and adaptation. A comprehensive multi-omics investigation of the BLA, spanning multiple cohorts, platforms, and analyses, revealed alterations in microglia and the extracellular matrix (ECM). Follow-up studies indicated that ER decreased microglia size, complexity, and phagocytosis, suggesting reduced immune surveillance. Loss also substantially increased ECM coverage, specifically targeting perineuronal nets surrounding parvalbumin interneurons, suggesting decreased plasticity and increased inhibition within the BLA following loss. Behavioral analyses suggest that these molecular effects are linked to impaired BLA salience evaluation, leading to a mismatch between stimulus and reaction intensity. These loss-like behaviors could be rescued by depleting BLA ECM during the removal period, helping us understand the mechanisms underlying loss and revealing novel molecular targets to ameliorate its impact., (© 2023. The Author(s).)

Fear and extinction learning are thought to generate distinct and competing memory representations in the hippocampus. How these memory representations modulate the expression of appropriate behavioral responses remains unclear. To investigate this question, we used cholera toxin B subunit to retrolabel ventral hippocampal (vHPC) neurons projecting to the infralimbic cortex (IL) and basolateral amygdala (BLA) and then quantified c-Fos immediate early gene activity within these populations following expression of either contextual fear recall or contextual fear extinction recall. Fear recall was associated with increased c-Fos expression in vHPC projections to the BLA, whereas extinction recall was associated with increased activity in vHPC projections to IL. A control experiment was performed to confirm that the apparent shift in projection neuron activity was associated with extinction learning rather than mere context exposure. Overall, results indicate that hippocampal contextual fear and extinction memory representations differentially activate vHPC projections to IL and BLA. These findings suggest that hippocampal memory representations orchestrate appropriate behavioral responses through selective activation of projection pathways., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Inc.)

Basolateral amygdala (BLA) projects widely across the macaque frontal cortex, and amygdalo-frontal projections are critical for appropriate emotional responding and decision making. While it is appreciated that single BLA neurons branch and project to multiple areas in frontal cortex, the organization and frequency of this branching has yet to be fully characterized. Here, we determined the projection patterns of more than 3,000 macaque BLA neurons. We found that one-third of BLA neurons had two or more distinct projection targets in frontal cortex and subcortical structures. The patterns of single BLA neuron projections to multiple areas were organized into repeating motifs that targeted distinct sets of areas in medial and ventral frontal cortex, indicative of separable BLA networks. Our findings begin to reveal the rich structure of single-neuron connections in the non-human primate brain, providing a neuroanatomical basis for the role of BLA in coordinating brain-wide responses to valent stimuli., Competing Interests: Declaration of interests The authors have no disclosures to report., (Copyright © 2023 Elsevier Inc. All rights reserved.)

It is widely accepted that Pavlovian fear conditioning requires activation of NMDA receptors (NMDARs) in the basolateral amygdala complex (BLA). However, it was recently shown that activation of NMDAR in the BLA is only required for fear conditioning when danger occurs unexpectedly; it is not required for fear conditioning when danger occurs as expected. This study tested the hypothesis that NMDARs in the BLA are engaged for Pavlovian fear conditioning when an animal's predictions regarding danger are in error. In each experiment, rats (females in Experiment 1 and males in Experiments 2-5) were conditioned to fear one stimulus, S1, when it was paired with foot-shock (S1→shock), and 48 h later, a second stimulus, S2, when it was presented in sequence with the already-conditioned S1 and foot-shock (S2→S1→shock). Conditioning to S2 occurred under a BLA infusion of the NMDAR antagonist, D-AP5 or vehicle. The subsequent tests of freezing to S2 alone and S1 alone revealed that the antagonist had no effect on conditioning to S2 when the shock occurred exactly as predicted by the S1, but disrupted this conditioning when the shock occurred earlier/later than predicted by S1, or at a stronger/weaker intensity. These results imply that errors in the timing or intensity of a predicted foot-shock engage NMDARs in the BLA for Pavlovian fear conditioning. They are discussed in relation to theories which propose a role for prediction error in determining how experiences are organized in memory and how activation of NMDAR in the BLA might contribute to this organization. SIGNIFICANCE STATEMENT This study is significant in showing that prediction error determines how a new experience is encoded with respect to a past experience and, thereby, whether NMDA receptors (NMDARs) in the basolateral amygdala complex (BLA) encode the new experience. When prediction error is small (e.g., danger occurs as and when expected), the new experience is encoded together with a past experience as part of the same "mental model," and NMDAR activation in the BLA is not needed for this encoding. By contrast, when prediction error is large (e.g., danger occurs at an unexpected intensity or time), the new experience is encoded separately from the past experience as part of a new mental model, and NMDAR activation in the BLA is needed for this encoding., (Copyright © 2023 the authors.)

Vicarious reward, essential to social learning and decision making, is theorized to engage select brain regions similarly to experienced reward to generate a shared experience. However, it is just as important for neural systems to also differentiate vicarious from experienced rewards for social interaction. Here, we investigated the neuronal interaction between the primate anterior cingulate cortex gyrus (ACCg) and the basolateral amygdala (BLA) when social choices made by monkeys led to either vicarious or experienced reward. Coherence between ACCg spikes and BLA local field potential (LFP) selectively increased in gamma frequencies for vicarious reward, whereas it selectively increased in alpha/beta frequencies for experienced reward. These respectively enhanced couplings for vicarious and experienced rewards were uniquely observed following voluntary choices. Moreover, reward outcomes had consistently strong directional influences from ACCg to BLA. Our findings support a mechanism of vicarious reward where social agency is tagged by interareal coordination frequency within the same shared pathway., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

One of the most sought-after topics in neuroscience is to understand how the environment regulates the activity and function of neural circuitry and subsequently influences relevant behaviors. In response to alterations in the environment, the neural circuits undergo adaptive changes ranging from gene expression changes to altered cellular function. Performing sequencing of the transcriptome involved in these behavior-related circuits will provide clues to accurately dissect the detailed mechanisms of related behavior. Here, we describe methods for marking and collecting the ventral hippocampus-projecting basolateral amygdala neurons, which have been repeatedly implicated in regulation of anxiety-like behavior, and subsequently constructing a library ready for sequencing. Specifically, the reported approaches include adeno-associated virus injection, acute brain slice isolation, cell suspension preparation, cell extraction, and cDNA library construction. By utilizing the techniques described here, researchers can comprehensively investigate the transcriptional levels of neural clusters embedded in particular circuits and discover potential pathogenic and therapeutic targets for behavior-relevant disorders. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Tagging of behavior-related neural circuits Basic Protocol 2: Isolation and capture of fluorescent-positive cells Basic Protocol 3: Foundation of sequencing library., (© 2023 Wiley Periodicals LLC.)

Behavioral flexibility and timely reactions to salient stimuli are essential for survival. The subcortical thalamic-basolateral amygdala (BLA) pathway serves as a shortcut for salient stimuli ensuring rapid processing. Here, we show that BLA neuronal and thalamic axonal activity in mice mirror the defensive behavior evoked by an innate visual threat as well as an auditory learned threat. Importantly, perturbing this pathway compromises defensive responses to both forms of threats, in that animals fail to switch from exploratory to defensive behavior. Despite the shared pathway between the two forms of threat processing, we observed noticeable differences. Blocking β-adrenergic receptors impairs the defensive response to the innate but not the learned threats. This reduced defensive response, surprisingly, is reflected in the suppression of the activity exclusively in the BLA as the thalamic input response remains intact. Our side-by-side examination highlights the similarities and differences between innate and learned threat-processing, thus providing new fundamental insights., Competing Interests: VK, IF, NM, PK, KY, SN No competing interests declared, (© 2023, Khalil, Faress, Mermet-Joret et al.)

Dysregulated fear reactions can result from maladaptive processing of trauma-related memories. In post-traumatic stress disorder (PTSD) and other psychiatric disorders, dysfunctional extinction learning prevents discretization of trauma-related memory engrams and generalizes fear responses. Although PTSD may be viewed as a memory-based disorder, no approved treatments target pathological fear memory processing. Hippocampal sharp wave-ripples (SWRs) and concurrent neocortical oscillations are scaffolds to consolidate contextual memory, but their role during fear processing remains poorly understood. Here, we show that closed-loop, SWR triggered neuromodulation of the medial forebrain bundle (MFB) can enhance fear extinction consolidation in male rats. The modified fear memories became resistant to induced recall (i.e., 'renewal' and 'reinstatement') and did not reemerge spontaneously. These effects were mediated by D2 receptor signaling-induced synaptic remodeling in the basolateral amygdala. Our results demonstrate that SWR-triggered closed-loop stimulation of the MFB reward system enhances extinction of fearful memories and reducing fear expression across different contexts and preventing excessive and persistent fear responses. These findings highlight the potential of neuromodulation to augment extinction learning and provide a new avenue to develop treatments for anxiety disorders., (© 2023. The Author(s).)

Astrocytes are key cellular regulators within the brain. The basolateral amygdala (BLA) is implicated in fear memory processing, yet most research has entirely focused on neuronal mechanisms, despite a significant body of work implicating astrocytes in learning and memory. In the present study, we used in vivo fiber photometry in C57BL/6J male mice to record from amygdalar astrocytes across fear learning, recall, and three separate periods of extinction. We found that BLA astrocytes robustly responded to foot shock during acquisition, their activity remained remarkably elevated across days in comparison to unshocked control animals, and their increased activity persisted throughout extinction. Further, we found that astrocytes responded to the initiation and termination of freezing bouts during contextual fear conditioning and recall, and this behavior-locked pattern of activity did not persist throughout the extinction sessions. Importantly, astrocytes do not display these changes while exploring a novel context, suggesting that these observations are specific to the original fear-associated environment. Chemogenetic inhibition of fear ensembles in the BLA did not affect freezing behavior or astrocytic calcium dynamics. Overall, our work presents a real-time role for amygdalar astrocytes in fear processing and provides new insight into the emerging role of these cells in cognition and behavior. SIGNIFICANCE STATEMENT We show that basolateral amygdala astrocytes are robustly responsive to negative experiences, like shock, and display changed calcium activity patterns through fear learning and memory. Additionally, astrocytic calcium responses become time locked to the initiation and termination of freezing behavior during fear learning and recall. We find that astrocytes display calcium dynamics unique to a fear-conditioned context, and chemogenetic inhibition of BLA fear ensembles does not have an impact on freezing behavior or calcium dynamics. These findings show that astrocytes play a key real-time role in fear learning and memory., (Copyright © 2023 the authors.)

Prefrontal cortical maturation coincides with adolescent transitions in social engagement, suggesting that it influences social development. The anterior cingulate cortex (ACC) is important for social interaction, including ACC outputs to the basolateral amygdala (BLA). However, little is known about ACC-BLA sensitivity to the social environment and if this changes during maturation. Here, we used brief (2-hour) isolation to test the immediate impact of changing the social environment on the ACC-BLA circuit and subsequent shifts in social behavior of adolescent and adult rats. We found that optogenetic inhibition of the ACC during brief isolation reduced isolation-driven facilitation of social interaction across ages. Isolation increased activity of ACC-BLA neurons across ages, but altered the influence of ACC on BLA activity in an age-dependent manner. Isolation reduced the inhibitory impact of ACC stimulation on BLA neurons in a frequency-dependent manner in adults, but uniformly suppressed ACC-driven BLA activity in adolescents. This work identifies isolation-driven alterations in an ACC-BLA circuit, and the ACC itself as an essential region sensitive to social environment and regulates its impact on social behavior in both adults and adolescents., (© The Author(s) 2023. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Increasing evidence has connected the development of certain neuropsychiatric disorders, as well as neurodegenerative diseases, to stress-induced dysregulation of the immune system. We have shown that escapable (ES) and inescapable (IS) footshock stress, and memories associated with ES or IS, can differentially alter inflammatory-related gene expression in brain in a region dependent manner. We have also demonstrated that the basolateral amygdala (BLA) regulates stress- and fear memory-induced alterations in sleep, and that differential sleep and immune responses in the brain to ES and IS appear to be integrated during fear conditioning and then reproduced by fear memory recall. In this study, we investigated the role of BLA in influencing regional inflammatory responses within the hippocampus (HPC) and medial prefrontal cortex (mPFC) by optogenetically stimulating or inhibiting BLA in male C57BL/6 mice during footshock stress in our yoked shuttlebox paradigm based on ES and IS. Then, mice were immediately euthanized and RNA extracted from brain regions of interest and loaded into NanoString® Mouse Neuroinflammation Panels for compilation of gene expression profiles. Results showed differential regional effects in gene expression and activated pathways involved in inflammatory-related signaling following ES and IS, and these differences were altered depending on amygdalar excitation or inhibition. These findings demonstrate that the stress-induced immune response, or "parainflammation", is affected by stressor controllability and that BLA influences regional parainflammation to ES or IS in HPC and mPFC. The study illustrates how stress-induced parainflammation can be regulated at the neurocircuit level and suggests that this approach can be useful for uncovering circuit and immune interactions in mediating differential stress outcomes., (© 2023. The Author(s).)

Introduction: Threatening environmental cues often generate enduring fear memories, but how these are formed and stored remains actively investigated. Recall of a recent fear memory is thought to reflect reactivation of neurons, in multiple brain regions, activated during memory formation, indicating that anatomically distributed and interconnected neuronal ensembles comprise fear memory engrams. The extent to which anatomically specific activation-reactivation engrams persist during long-term fear memory recall, however, remains largely unexplored. We hypothesized that principal neurons in the anterior basolateral amygdala (aBLA), which encode negative valence, acutely reactivate during remote fear memory recall to drive fear behavior., Methods: Using adult offspring of TRAP2 and Ai14 mice, persistent tdTomato expression was used to "TRAP" aBLA neurons that underwent Fos-activation during contextual fear conditioning (electric shocks) or context only conditioning (no shocks) ( n = 5/group). Three weeks later, mice were re-exposed to the same context cues for remote memory recall, then sacrificed for Fos immunohistochemistry., Results: TRAPed (tdTomato +), Fos +, and reactivated (double-labeled) neuronal ensembles were larger in fear- than context-conditioned mice, with the middle sub-region and middle/caudal dorsomedial quadrants of aBLA displaying the greatest densities of all three ensemble populations. Whereas tdTomato + ensembles were dominantly glutamatergic in context and fear groups, freezing behavior during remote memory recall was not correlated with ensemble sizes in either group., Discussion: We conclude that although an aBLA-inclusive fear memory engram forms and persists at a remote time point, plasticity impacting electrophysiological responses of engram neurons, not their population size, encodes fear memory and drives behavioral manifestations of long-term fear memory recall., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Hammack, Fischer, Andrade and Toney.)