A 'double-edged' role for type-5 metabotropic glutamate receptors in pain disclosed by light-sensitive drugs.

We used light-sensitive drugs to identify the brain region-specific role of mGlu5 metabotropic glutamate receptors in the control of pain. Optical activation of systemic JF-NP-26, a caged, normally inactive, negative allosteric modulator (NAM) of mGlu5 receptors, in cingulate, prelimbic, and infralimbic cortices and thalamus inhibited neuropathic pain hypersensitivity. Systemic treatment of alloswitch-1, an intrinsically active mGlu5 receptor NAM, caused analgesia, and the effect was reversed by light-induced drug inactivation in the prelimbic and infralimbic cortices, and thalamus. This demonstrates that mGlu5 receptor blockade in the medial prefrontal cortex and thalamus is both sufficient and necessary for the analgesic activity of mGlu5 receptor antagonists. Surprisingly, when the light was delivered in the basolateral amygdala, local activation of systemic JF-NP-26 reduced pain thresholds, whereas inactivation of alloswitch-1 enhanced analgesia. Electrophysiological analysis showed that alloswitch-1 increased excitatory synaptic responses in prelimbic pyramidal neurons evoked by stimulation of presumed BLA input, and decreased BLA-driven feedforward inhibition of amygdala output neurons. Both effects were reversed by optical silencing and reinstated by optical reactivation of alloswitch-1. These findings demonstrate for the first time that the action of mGlu5 receptors in the pain neuraxis is not homogenous, and suggest that blockade of mGlu5 receptors in the BLA may limit the overall analgesic activity of mGlu5 receptor antagonists. This could explain the suboptimal effect of mGlu5 NAMs on pain in human studies and validate photopharmacology as an important tool to determine ideal target sites for systemic drugs.

Ketamine ameliorates post-traumatic social avoidance by erasing the traumatic memory encoded in VTA-innervated BLA engram cells.

Erasing traumatic memory during memory reconsolidation is a promising retrieval-extinction strategy for post-traumatic stress disorder (PTSD). Here, we developed an acute social defeat stress (SDS) mouse model with short-term and re-exposure-evoked long-term social avoidance. SDS-associated traumatic memories were identified to be stored in basolateral amygdala (BLA) engram cells. A single intraperitoneal administration of subanesthetic-dose ketamine within, but not beyond, the re-exposure time window significantly alleviates SDS-induced social avoidance, which reduces the activity and quantity of reactivated BLA engram cells. Furthermore, activation or inhibition of dopaminergic projections from the ventral tegmental area to the BLA effectively mimics or blocks the therapeutic effect of re-exposure with ketamine and is dopamine D2 receptor dependent. Single-cell RNA sequencing reveals that re-exposure with ketamine triggered significant changes in memory-related pathways in the BLA. Together, our research advances the understanding of how ketamine mitigates PTSD symptoms and offers promising avenues for developing more effective treatments for trauma-related disorders.

Activation of NPY Receptors in the BLA Inhibits Projections to the Bed Nucleus of the Stria Terminalis and Buffers Stress-Induced Decreases in Social Interaction in Male Rats.

Neuropeptide Y (NPY) increases resilience and buffers behavioral stress responses in male rats in part through decreasing the excitability of principal output neurons in the basolateral amygdala (BLA). Intra-BLA administration of NPY acutely increases social interaction (SI) through activation of either Y1 or Y5 receptors, whereas repeated NPY (rpNPY) injections (once daily for 5 d) produce persistent increases in SI through Y5 receptor-mediated neuroplasticity in the BLA. In this series of studies, we characterized the neural circuits from the BLA that underlie these behavioral responses to NPY. Using neuronal tract tracing, NPY Y1 and Y5 receptor immunoreactivity was identified on subpopulations of BLA neurons projecting to the bed nucleus of the stria terminalis (BNST) and the central nucleus of the amygdala (CeA). Inhibition of BLA→BNST, but not BLA→CeA, neurons using projection-restricted, cre-driven designer receptors exclusively activated by designer drug-Gi expression increased SI and prevented stress-induced decreases in SI produced by a 30 min restraint stress. This behavioral profile was similar to that seen after both acute and rpNPY injections into the BLA. Intracellular recordings of BLA→BNST neurons demonstrated NPY-mediated inhibition via suppression of H currents, as seen previously. Repeated intra-BLA injections of NPY, which are associated with the induction of BLA neuroplasticity, decreased the activity of BLA→BNST neurons and decreased their dendritic complexity. These results demonstrate that NPY modulates the activity of BNST-projecting BLA neurons, suggesting that this pathway contributes to the stress-buffering actions of NPY and provides a novel substrate for the proresilient effects of NPY.

The modulatory role of serotonin-1A receptors of the basolateral amygdala and dorsal periaqueductal gray on the impact of hormonal variation on the conditioned fear response.

Despite significant advances in the study of fear and fear memory formation, little is known about fear learning and expression in females. This omission has been proven surprising, as normal and pathological behaviors are highly influenced by ovarian hormones, particularly estradiol and progesterone. In the current study, we investigated the joint influence of serotonin (5-HT) neurotransmission and estrous cycle phases (low or high levels of estradiol and progesterone) on the expression of conditioned fear in a group of female rats that were previously divided according to their response to stressful stimuli into low or high anxiety-like subjects. The baseline amplitude of the unconditioned acoustic startle responses was high in high-anxiety female rats, with no effect on the estrous cycle observed. Data collected during the proestrus-estrus phase revealed that low-anxiety rats had startle amplitudes similar to those of high-anxiety rats. It is supposed that high-anxiety female rats benefit from increased estradiol and progesterone levels to achieve comparable potentiated startle amplitudes. In contrast, female rats experienced a significant decrease in hormone levels during the Diestrus phase. This decrease is believed to play a role in preventing them from displaying a heightened startle response when faced with strongly aversive stimuli. Data collected after 5-HT and 8-OH-DPAT were administered into the basolateral nuclei and dorsal periaqueductal gray suggest that 5-HT neurotransmission works with progesterone and estrogen to reduce startle potentiation, most likely by activating the serotonin-1A receptor subtype.

Estrogen receptor beta signaling enhances extinction memory recall for heroin-conditioned cues in a sex- and region-specific manner.

Return to use, or relapse, is a major challenge in the treatment of opioid use disorder (OUD). Relapse can be precipitated by several factors, including exposure to drug-conditioned cues. Identifying successful treatments to mitigate cue-induced relapse has been challenging, perhaps due to extinction memory recall (EMR) deficits. Previously, inhibition of estradiol (E2) signaling in the basolateral amygdala (BLA) impaired heroin-cue EMR. This effect was recapitulated by antagonism of BLA estrogen receptors (ER) in a sex-specific manner such that blocking ERα in males, but ERß in females, impaired EMR. However, it is unclear whether increased E2 signaling, in the BLA or systemically, enhances heroin-cue EMR. We hypothesized that ERß agonism would enhance heroin-cue EMR in a sex- and region-specific manner. To determine the capacity of E2 signaling to improve EMR, we pharmacologically manipulated ERß across several translationally designed experiments. First, male and female rats acquired heroin or sucrose self-administration. Next, during a cued extinction session, we administered diarylpropionitrile (DPN, an ERß agonist) and tested anxiety-like behavior on an open field. Subsequently, we assessed EMR in a cue-induced reinstatement test and, finally, measured ERß expression in several brain regions. Across all experiments, females took more heroin and sucrose than males and had greater responses during heroin-cued extinction. Administration of DPN in the BLA enhanced EMR in females only, driven by ERß's impacts on memory consolidation. Interestingly, however, systemic DPN administration improved EMR for heroin cues in both sexes across several different tests, but did not impact sucrose-cue EMR. Immunohistochemical analysis of ERß expression across several different brain regions showed that females only had greater expression of ERß in the basal nucleus of the BLA. Here, in several preclinical experiments, we demonstrated that ERß agonism enhances heroin-cue EMR and has potential utility in combatting cue-induced relapse.

The dose-dependent effect of the D2R agonist quinpirole microinjected into the ventral pallidum on information flow in the limbic system.

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 (D2R) agonist quinpirole dose-dependently facilitates memory consolidation in inhibitory avoidance and spatial learning. In the VP, D2R 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 D2Rs in the regulation of limbic information flow.

Scopolamine infusion in the basolateral amygdala after saccharin intake induces conditioned taste avoidance in rats.

RATIONALE: Muscarinic receptor activity in the basolateral amygdala (BLA) is known to be involved in plasticity mechanisms that underlie emotional learning. The BLA is involved in the Attenuation of Neophobia, an incidental taste learning task in which a novel taste becomes familiar and recognized as safe. OBJECTIVE: Here we assessed the role of muscarinic receptor activity in the BLA in incidental taste learning. METHODS: Young adult male Wistar rats were bilaterally implanted with cannulas aimed at BLA. After recovery, rats were randomly assigned to either vehicle or muscarinic antagonist group, for each experiment. We tested the effect of specific and non-specific muscarinic antagonists administered either 1) 20 min before novel taste presentation; 2) immediately after novel taste presentation; 3) immediately after retrieval (the second taste presentation on Day 5 -S2-) or immediately after the fifth taste presentation on Day 8 (S5). RESULTS: Non-specific muscarinic receptor antagonist scopolamine infused prior to novel taste, while not affecting novel taste preference, abolished AN, i.e., the increased preference observed in control animals on the second presentation. When administered after taste consumption, intra-BLA scopolamine not only prevented AN but caused a steep decrease in the taste preference on the second presentation. This scopolamine-induced taste avoidance was not dependent on taste novelty, nor did it generalize to another novel taste. Targeting putative postsynaptic muscarinic receptors with specific M1 or M3 antagonists appeared to produce a partial taste avoidance, while M2 antagonism had no effect. CONCLUSION: These data suggest that if a salient gustatory experience is followed by muscarinic receptors antagonism in the BLA, it will be strongly and persistently avoided in the future. The study also shows that scopolamine is not just an amnesic drug, and its cognitive effects may be highly dependent on the task and the structure involved.

Blockade of pre- and post-synaptic GABAB receptors in the anteroventral bed nucleus of stria terminalis produces anxiolytic-like and anxiety-like effects in parkinsonian rats respectively.

The anteroventral bed nucleus of stria terminalis (avBNST) is a limbic forebrain region involved in the regulation of anxiety, and expresses GABAB receptors, which are located at both pre- and post-synaptic sites. However, it is unclear how blockade of these receptors affects anxiety-like behaviors, particularly in Parkinson's disease (PD)-related anxiety. In the present study, unilateral 6-hydroxydopamine (6-OHDA) lesions of the substantia nigra pars compacta in rats induced anxiety-like behaviors, and increased GABA release and decreased glutamate release in the avBNST, as well as decreased level of dopamine (DA) in the basolateral amygdala (BLA). Intra-avBNST injection of pre-synaptic GABAB receptor antagonist CGP36216 produced anxiolytic-like effects, while the injection of post-synaptic GABAB receptor antagonist CGP35348 induced anxiety-like responses in both sham and 6-OHDA rats. Intra-avBNST injection of CGP36216 inhibited the GABAergic neurons and increased GABA/glutamate ratio in the avBNST and increased levels of DA and serotonin (5-HT) in the BLA; conversely, CGP35348 produced opposite effects on the firing activity of avBNST GABAergic neurons and levels of the neurotransmitters in the avBNST and BLA. Moreover, the doses of the antagonists producing significant behavioral effects in 6-OHDA rats were lower than those in sham rats, and the duration of action of the antagonists on the firing rate of the neurons and release of the neurotransmitters was prolonged in 6-OHDA rats. Altogether, these findings suggest that pre- and post-synaptic GABAB receptors in the avBNST are implicated in PD-related anxiety-like behaviors, and degeneration of the nigrostriatal pathway enhances functions and/or upregulates expression of these receptors.

Effects of acute inhibition of dopamine ß-hydroxylase on neural responses to pups in adult virgin male California mice (Peromyscus californicus).

The neural mechanisms underlying paternal care in biparental mammals are not well understood. The California mouse (Peromyscus californicus) is a biparental rodent in which virtually all fathers are attracted to pups, while virgin males vary widely in their behavior toward unrelated infants, ranging from attacking to avoiding to huddling and grooming pups. We previously showed that pharmacologically inhibiting the synthesis of the neurotransmitter norepinephrine (NE) with the dopamine ß-hydroxylase inhibitor nepicastat reduced the propensity of virgin male and female California mice to interact with pups. The current study tested the hypothesis that nepicastat would reduce pup-induced c-Fos immunoreactivity, a cellular marker of neural activity, in the medial preoptic area (MPOA), medial amygdala (MeA), basolateral amygdala (BLA), and bed nucleus of the stria terminalis (BNST), brain regions implicated in the control of parental behavior and/or anxiety. Virgin males were injected with nepicastat (75 mg/kg, i.p.) or vehicle 2 hours prior to exposure to either an unrelated pup or novel object for 60 minutes (n = 4-6 mice per group). Immediately following the 60-minute stimulus exposure, mice were euthanized and their brains were collected for c-Fos immunohistochemistry. Nepicastat reduced c-Fos expression in the MeA and MPOA of pup-exposed virgin males compared to vehicle-injected controls. In contrast, nepicastat did not alter c-Fos expression in any of the above brain regions following exposure to a novel object. Overall, these results suggest that the noradrenergic system might influence MeA and MPOA function to promote behavioral interactions with pups in virgin males.

Planar cell polarity proteins mediate ketamine-induced restoration of glutamatergic synapses in prefrontal cortical neurons in a mouse model for chronic stress.

Single administration of low-dose ketamine has both acute and sustained anti-depressant effects. Sustained effect is associated with restoration of glutamatergic synapses in medial prefrontal cortic (mFPC) neurons. Ketamine induced profound changes in a number of molecular pathways in a mouse model for chronic stress. Cell-cell communication analyses predicted that planar-cell-polarity (PCP) signaling was decreased after chronic administration of corticosterone but increased following ketamine administration in most of the excitatory neurons. Similar decrease of PCP signaling in excitatory neurons was predicted in dorsolateral prefrontal cortical (dl-PFC) neurons of patients with major depressive disorder (MDD). We showed that the basolateral amygdala (BLA)-projecting infralimbic prefrontal cortex (IL PFC) neurons regulate immobility time in the tail suspension test and food consumption. Conditionally knocking out Celsr2 and Celsr3 or Prickle2 in the BLA-projecting IL PFC neurons abolished ketamine-induced synapse restoration and behavioral remission. Therefore, PCP proteins in IL PFC-BLA neurons mediate synapse restoration induced by of low-dose ketamine.

Predatory Odor Exposure as a Potential Paradigm for Studying Emotional Modulation of Memory Consolidation-The Role of the Noradrenergic Transmission in the Basolateral Amygdala.

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.

Dorsal raphe nucleus to basolateral amygdala 5-HTergic neural circuit modulates restoration of consciousness during sevoflurane anesthesia.

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.

Formation of chronic morphine withdrawal memories requires C1QL3-mediated regulation of PSD95 in the mouse basolateral amygdala.

Chronic morphine withdrawal memory formation is a complex process influenced by various molecular mechanisms. In this study, we aimed to investigate the contributions of the basolateral amygdala (BLA) and complement component 1, q subcomponent-like 3 (C1QL3), a secreted and presynaptically targeted protein, to the formation of chronic morphine (repeat dosing of morphine) withdrawal memory using conditioned place aversion (CPA) and chemogenetic methods. We conducted experiments involving the inhibition of the BLA during naloxone-induced withdrawal to assess its impact on CPA scores, providing insights into the significance of the BLA in the chronic morphine memory formation process. We also examined changes in C1ql3/C1QL3 expression within the BLA following conditioning. Immunofluorescence analysis revealed the colocalization of C1QL3 and the G protein-coupled receptor, brain-specific angiogenesis inhibitor 3 (BAI3) in the BLA, supporting their involvement in synaptic development. Moreover, we downregulated C1QL3 expression in the BLA to investigate its role in chronic morphine withdrawal memory formation. Our findings revealed that BLA inhibition during naloxone-induced withdrawal led to a significant reduction in CPA scores, confirming the critical role of the BLA in this memory process. Additionally, the upregulation of C1ql3 expression within the BLA postconditioning suggested its participation in withdrawal memory formation. The colocalization of C1QL3 and BAI3 in the BLA further supported their involvement in synaptic development. Furthermore, downregulation of C1QL3 in the BLA effectively hindered chronic morphine withdrawal memory formation, emphasizing its pivotal role in this process. Notably, we identified postsynaptic density protein 95 (PSD95) as a potential downstream effector of C1QL3 during chronic morphine withdrawal memory formation. Blocking PSD95 led to a significant reduction in the CPA score, and it appeared that C1QL3 modulated the ubiquitination-mediated degradation of PSD95, resulting in decreased PSD95 protein levels. This study underscores the importance of the BLA, C1QL3 and PSD95 in chronic morphine withdrawal memory formation. It provides valuable insights into the underlying molecular mechanisms, emphasizing their significance in this intricate process.

Urolithin A Inhibits Anterior Basolateral Amygdala to Ventral Hippocampal CA1 Circuit to Ameliorate Amyloid-ß-Impaired Social Ability.

Background: Anxiety and social withdrawal are highly prevalent among patients with Alzheimer's disease (AD). However, the neural circuit mechanisms underlying these symptoms remain elusive, and there is a need for effective prevention strategies. Objective: This study aims to elucidate the neural circuitry mechanisms underlying social anxiety in AD. Methods: We utilized 5xFAD mice and conducted a series of experiments including optogenetic manipulation, Tandem Mass Tag-labeled proteome analysis, behavioral assessments, and immunofluorescence staining. Results: In 5xFAD mice, we observed significant amyloid-ß (Aß) accumulation in the anterior part of basolateral amygdala (aBLA). Behaviorally, 6-month-old 5xFAD mice displayed excessive social avoidance during social interaction. Concurrently, the pathway from aBLA to ventral hippocampal CA1 (vCA1) was significantly activated and exhibited a disorganized firing patterns during social interaction. By optogenetically inhibiting the aBLA-vCA1 pathway, we effectively improved the social ability of 5xFAD mice. In the presence of Aß accumulation, we identified distinct changes in the protein network within the aBLA. Following one month of administration of Urolithin A (UA), we observed significant restoration of the abnormal protein network within the aBLA. UA treatment also attenuated the disorganized firings of the aBLA-vCA1 pathway, leading to an improvement in social ability. Conclusions: The aBLA-vCA1 circuit is a vulnerable pathway in response to Aß accumulation during the progression of AD and plays a crucial role in Aß-induced social anxiety. Targeting the aBLA-vCA1 circuit and UA administration are both effective strategies for improving the Aß-impaired social ability.

Terminalia chebula attenuates restraint stress-induced memory impairment and synaptic loss in the dentate gyrus of the hippocampus and the basolateral and central nuclei of the amygdala by inhibiting oxidative damage.

Chronic restraint stress induces cognitive abnormalities through changes in synapses and oxidant levels in the amygdala and hippocampus. Given the neuroprotective effects of fruit of Terminalia chebula (Halileh) in different experimental models, the present investigation aimed to address whether Terminalia chebula is able to reduce chronic restraint stress-induced behavioral, synaptic and oxidant markers in the rat model. Thirty-two male Wistar rats were randomly divided into four groups as follows: control (did not receive any treatment and were not exposed to stress), stress (restraint stress for 2 h a day for 14 consecutive days), Terminalia chebula (received 200 mg/kg hydroalcoholic extract of Terminalia chebula), and stress + Terminalia chebula groups (received 200 mg/kg extract of Terminalia chebula twenty minutes before stress) (n = 8 in each group). We used the shuttle box test to assess learning and memory, Golgi-Cox staining to examine dendritic spine density in the dentate gyrus region of the hippocampus and the basolateral and central nuclei of the amygdala, and total antioxidant capacity (TAC) and total oxidant status (TOS) in the brain. The shuttle box test results demonstrated that Terminalia chebula treatment had a profound positive effect on memory parameters, including step-through latency (STL) and time spent in the dark room, when compared to the stress group. Daily oral treatment with Terminalia chebula effectively suppressed the loss of neural spine density in the dentate gyrus region of the hippocampus and the basolateral and central nuclei of the amygdala caused by chronic restraint stress, as demonstrated by Golgi-Cox staining. Additionally, the results indicate that Terminalia chebula significantly reduced the TOS and increased TAC in the brain compared to the stress group. In conclusion, our results suggest that Terminalia chebula improved memory impairment and synaptic loss in the dentate gyrus of the hippocampus and the basolateral and central nuclei of the amygdala induced by restraint stress via inhibiting oxidative damage.

The impact of chronic fluoxetine treatment in adolescence or adulthood on context fear memory and perineuronal nets.

Selective serotonin reuptake inhibitors, such as fluoxetine (Prozac), are commonly prescribed pharmacotherapies for anxiety. Fluoxetine may be a useful adjunct because it can reduce the expression of learned fear in adult rodents. This effect is associated with altered expression of perineuronal nets (PNNs) in the amygdala and hippocampus, two brain regions that regulate fear. However, it is unknown whether fluoxetine has similar effects in adolescents. Here, we investigated the effect of fluoxetine exposure during adolescence or adulthood on context fear memory and PNNs in the basolateral amygdala (BLA), the CA1 subregion of the hippocampus, and the medial prefrontal cortex in rats. Fluoxetine impaired context fear memory in adults but not in adolescents. Further, fluoxetine increased the number of parvalbumin (PV)-expressing neurons surrounded by a PNN in the BLA and CA1, but not in the medial prefrontal cortex, at both ages. Contrary to previous reports, fluoxetine did not shift the percentage of PNNs toward non-PV cells in either the BLA or CA1 in the adults, or adolescents. These findings demonstrate that fluoxetine differentially affects fear memory in adolescent and adult rats but does not appear to have age-specific effects on PNNs.

Role of amygdala astrocytes in different phases of contextual fear memory.

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).

Acetylcholine Engages Distinct Amygdala Microcircuits to Gate Internal Theta Rhythm.

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.

Restoration of nNOS Expression Rescues Autistic-Like Phenotypes Through Normalization of AMPA Receptor-Mediated Neurotransmission.

Autism spectrum disorder (ASD) is associated with a range of abnormalities characterized by deficits in socialization, communication, repetitive behaviors, and restricted interests. We have recently shown that neuronal nitric oxide synthase (nNOS) expression was decreased in the basolateral amygdala (BLA) of mice after postnatal valproic acid exposure. Neuronal activity-regulated pentraxin (Narp) could contribute to the regulation of the GluA4 2-amino-3-(5-methyl-3-oxo-1,2-oxazol-4-yl) propanoic acid (AMPA) subunits which are predominantly expressed in interneurons. However, the specific role of nNOS re-expression on excitatory neurotransmitter with relevance to ASD core symptoms in VPA-treated animals remains to be elucidated. Herein, nNOS overexpression using a lentiviral vector and L-arginine-activating PI3K-Akt-mTOR signaling can restore nNOS expression in the BLA induced by VPA. Restoration of nNOS expression in these mice was sufficient to reduce the severity of ASD-like behavioral patterns such that animals exhibited decreases in abnormal social interactions and communication, stereotyped/repetitive behaviors, and anxiety-like traits. Most strikingly, re-expression of nNOS upregulated surface expression of Narp and GluA4 in nNOS-positive interneuron as shown by immunoprecipitation and Western blotting. Whole-cell patch-clamp recordings demonstrated that restoration of nNOS had a significant enhancing effect on AMPA receptor-mediated excitatory glutamatergic synaptic neurotransmission, which was inhibited by disturbing the interaction between Narp and GluA4 in acutely dissociated BLA slices. Overall, these data offer a scientific basis for the additional study of nNOS re-expression as a promising therapeutic target by correcting AMPA receptor-mediated synaptic function in ASD and related neurodevelopmental disorders.

Memory consolidation drives the enhancement of remote cocaine memory via prefrontal circuit.

Remote memory usually decreases over time, whereas remote drug-cue associated memory exhibits enhancement, increasing the risk of relapse during abstinence. Memory system consolidation is a prerequisite for remote memory formation, but neurobiological underpinnings of the role of consolidation in the enhancement of remote drug memory are unclear. Here, we found that remote cocaine-cue associated memory was enhanced in rats that underwent self-administration training, together with a progressive increase in the response of prelimbic cortex (PrL) CaMKII neurons to cues. System consolidation was required for the enhancement of remote cocaine memory through PrL CaMKII neurons during the early period post-training. Furthermore, dendritic spine maturation in the PrL relied on the basolateral amygdala (BLA) input during the early period of consolidation, contributing to remote memory enhancement. These findings indicate that memory consolidation drives the enhancement of remote cocaine memory through a time-dependent increase in activity and maturation of PrL CaMKII neurons receiving a sustained BLA input.