Anxiety-like Behavior In Mice Research Articles

The endocannabinoid system is involved in the anxiety-like behavior induced by dual-frequency 2.65/0.8 GHz electromagnetic radiation in mice

As wireless communication devices gain popularity, concerns about the potential risks of environmental exposure to complex frequency electromagnetic radiation (EMR) on mental health have become a public health issue. Historically, EMR research has predominantly focused on single- frequency electromagnetic waves, neglecting the study of multi-frequency electromagnetic waves, which more accurately represent everyday life. To address these concerns, our study compared the emotional effects of single-frequency and dual-frequency EMR while exploring potential molecular mechanisms and intervention targets. Our results revealed that single-frequency EMR at 2.65 or 0.8 GHz did not induce anxiety-like behavior in mice. However, exposure to dual-frequency EMR at 2.65/0.8 GHz significantly led to anxiety-like behavior in mice. Further analysis of mouse sera revealed substantial increases in corticosterone and corticotrophin releasing hormone levels following exposure to 2.65/0.8 GHz EMR. Transcriptome sequencing indicated a significant decrease in the expression of Cnr1, encoding cannabinoid receptor 1 Type (CB1R), in the cerebral. This finding was consistently verified through western blot analysis, revealing a substantial reduction in CB1R content. Additionally, a significant decrease in the endocannabinoid 2-arachidonoylglycerol was observed in the cerebral cortex. Remarkably, administering the cannabinoid receptor agonist Win55-212-2 significantly alleviated the anxiety-like behavior, and the cannabinoid receptor antagonist AM251 effectively counteracted the anti-anxiety effects of Win55-212-2. In summary, our research confirmed that dual-frequency EMR is more likely to induce anxiety-like behavior in mice than single-frequency EMR, with implications for the hypothalamic–pituitary–adrenal axis and the endocannabinoid system. Furthermore, our findings suggest that Win55-212-2 may represent a novel avenue for researching and developing anti-EMR drugs.

Stress increases hepatic release of lipocalin 2 which contributes to anxiety-like behavior in mice.

Chronic stress induces anxiety disorders via both neural pathways and circulating factors. Although many studies have elucidated the neural circuits involved in stress-coping behaviors, the origin and regulatory mechanism of peripheral cytokines in behavioural regulation under stress conditions are not fully understood. Here, we identified a serum cytokine, lipocalin 2 (LCN2), that was upregulated in participants with anxiety disorders. Using a mouse model of chronic restraint stress (CRS), circulating LCN2 was found to be related to stress-induced anxiety-like behaviour via modulation of neural activity in the medial prefrontal cortex (mPFC). These results suggest that stress increases hepatic LCN2 via a neural pathway, leading to disrupted cortical functions and behaviour.

Intranasal delivery of shRNA to knockdown the 5HT-2A receptor enhances memory and alleviates anxiety

Short-hairpin RNAs (shRNA), targeting knockdown of specific genes, hold enormous promise for precision-based therapeutics to treat numerous neurodegenerative disorders. However, whether shRNA constructed molecules can modify neuronal circuits underlying certain behaviors has not been explored. We designed shRNA to knockdown the human HTR2A gene in vitro using iPSC-differentiated neurons. Multi-electrode array (MEA) results showed that the knockdown of the 5HT-2A mRNA and receptor protein led to a decrease in spontaneous electrical activity. In vivo, intranasal delivery of AAV9 vectors containing shRNA resulted in a decrease in anxiety-like behavior in mice and a significant improvement in memory in both mice (104%) and rats (92%) compared to vehicle-treated animals. Our demonstration of a non-invasive shRNA delivery platform that can bypass the blood–brain barrier has broad implications for treating numerous neurological mental disorders. Specifically, targeting the HTR2A gene presents a novel therapeutic approach for treating chronic anxiety and age-related cognitive decline.

Dexmedetomidine inhibits paraventricular corticotropin-releasing hormone neurons that attenuate acute stress-induced anxiety-like behavior in mice.

Dexmedetomidine has repeatedly shown to improve anxiety but the precise neural mechanisms underlying this effect remain incompletely understood. Here, we aim to explore the role of corticotropin-releasing hormone-producing hypothalamic paraventricular (CRHPVN) neurons in mediating the anxiolytic effects of dexmedetomidine. A social defeat stress mouse model was employed to evaluate the anxiolytic effects induced by dexmedetomidine through the elevated plus-maze, open-field test, and measurement of serum stress hormone levels. In vivo Ca2+ signal fibre photometry and ex vivo patch-clamp recordings were utilized to determine the excitability of CRHPVN neurons and investigate the specific mechanism involved. CRHPVN neuron modulation was achieved through chemogenetic activation or inhibition. Compared with saline, dexmedetomidine (40 µg/kg) alleviated anxiety-like behaviors. Additionally, dexmedetomidine reduced CRHPVN neuronal excitability. Chemogenetic activation of CRHPVN neurons decreased the time spent in the open arms of the elevated plus-maze and in the central area of the open-field test. Conversely, chemogenetic inhibition of CRHPVN neurons had the opposite effect. Moreover, the suppressive impact of dexmedetomidine on CRHPVN neurons was attenuated by the α2 receptor antagonist yohimbine. Our results indicate that the anxiety-like effects of dexmedetomidine are mediated via α2 adrenergic receptor-triggered inhibition of CRHPVN neuronal excitability in the hypothalamus.

SNX27: A trans-species cognitive modulator with implications for anxiety and stress susceptibility

Sorting Nexin 27 (SNX27) is a brain-enriched endosome-associated cargo adaptor that shapes excitatory control, being relevant for cognitive and reward processing, and for several neurological conditions. Despite this, SNX27's role in the nervous system remains poorly explored. To further understand SNX27 function, we performed an extensive behavioral characterization comprising motor, cognitive and emotional dimensions of SNX27+/− mice. Furthermore, attending on the recently described association between SNX27 function and cellular stress signaling mechanisms in vitro, we explored SNX27-stress interplay using a Caenorhabditis elegans Δsnx-27 mutant and wild-type (WT) rodents after stress exposure.SNX27+/− mice, as C. elegans Δsnx-27 mutants, present cognitive impairments, highlighting a conserved role for SNX27 in cognitive modulation across species. Interestingly, SNX27 downmodulation leads to anxiety-like behavior in mice evaluated in the Elevated Plus Maze (EPM). This anxious phenotype is associated with increased dendritic complexity of the bed nucleus of the stria terminalis (BNST) neurons, and increased complexity of the basolateral amygdala (BLA) pyramidal neurons. These findings highlight the still unknown role of SNX27 in anxiety regulation.Moreover, we uncovered a direct link between SNX27 dysfunction and stress susceptibility in C. elegans and found that stress-exposed rodents display decreased SNX27 levels in stress-susceptible brain regions.Altogether, we provided new insights on SNX27's relevance in anxiety-related behaviors and neuronal structure in stress-associated brain regions.

Polystyrene microplastics induce anxiety via HRAS derived PERK-NF-κB pathway

Exposure to environmentally hazardous substances is recognized as a significant risk factor for neurological associated disorders. Among these substances, polystyrene microplastics (PS-MPs), widely utilized in various consumer products, have been reported to exhibit neurotoxicity. However, the potential association of PS-MPs with abnormal anxiety behaviors, along with the underlying molecular mechanisms and key proteins involved, remains insufficiently explored. Here, we delineated the potential mechanisms of PS-MPs-induced anxiety through proteomics and molecular investigations. We characterized the PS-MPs, observed their accumulation in the brain, leading to anxiety-like behavior in mice, which is correlated with microglia activation and pro-inflammatory response. Consistent with these findings, our studies on BV2 microglia cells showed that PS-MPs activated NF-κB-mediated inflammation resulting in the upregulation of pro-inflammatory cytokines such as TNFα and IL-1β. Of particular significance, HRAS was identified as a key factor in the PS-MPs induced pro-inflammatory response through whole proteomics analysis, and knockdown of H-ras effectively inhibited PS-MPs induced PERK-NF-κB activation and associated pro-inflammatory response in microglia cells. Collectively, our findings highlight that PS-MPs induce anxiety of mice via the activation of the HRAS-derived PERK-NF-κB pathway in microlglia. Our results contribute valuable insights into the molecular mechanisms of PS-MPs-induced anxiety, and may offer implications for addressing neurotoxicity and prevention the adverse effects of environmentally hazardous substances, including microplastics.

Dulaglutide treatment reverses depression-like behavior and hippocampal metabolomic homeostasis in mice exposed to chronic mild stress.

Treatment strategies for depression based on interventions for glucose and lipid metabolism disorders are receiving increasing attention. Investigating the mechanism of their antidepressant effect and exploring new diagnostic and therapeutic biomarkers have attracted increasing attention. Dulaglutide, a long-acting GLP-1 receptor agonist, has been reported to alleviate cognitive deficits and neuronal damage. However, the antidepressant effect of dulaglutide and, especially, the underlying mechanism are still poorly understood. In this study, we aimed to explore the underlying biomarkers of depression and potential modulatory targets of dulaglutide in chronic mild stress (CMS) mice. Sixty mice were randomly divided into a control group (CON group), a CMS+Vehicle group (CMS+Veh group), a CMS+0.3mg/kg dulaglutide group (Low Dula group), and a CMS+0.6mg/kg dulaglutide group (High Dula group). Numerous behavioral tests, mainly the open field test, forced swimming test, and tail suspension test, were applied to evaluate the potential effect of dulaglutide treatment on anxiety- and depression-like behaviors in mice exposed to chronic stress. Furthermore, a liquid chromatography-tandem mass spectrometry-based metabolomics approach was utilized to investigate the associated mechanisms of dulaglutide treatment. Three weeks of dulaglutide treatment significantly reversed depressive-like but not anxiety-like behaviors in mice exposed to chronic stress for 4 weeks. The results from the metabolomics analysis showed that a total of 20 differentially expressed metabolites were identified between the CON and CMS+Veh groups, and 46 metabolites were selected between the CMS+Veh and High Dula groups in the hippocampus of the mice. Comprehensive analysis indicated that lipid metabolism, amino acid metabolism, energy metabolism, and tryptophan metabolism were disrupted in model mice that experienced depression and underwent dulaglutide therapy. The antidepressant effects of dulaglutide in a CMS depression model were confirmed. We identified 64 different metabolites and four major pathways associated with metabolic pathophysiological processes. These primary data provide a new perspective for understanding the antidepressant-like effects of dulaglutide and may facilitate the use of dulaglutide as a potential therapeutic strategy for depression.

Microglial activation in the lateral amygdala promotes anxiety-like behaviors in mice with chronic moderate noise exposure.

Long-term non-traumatic noise exposure, such as heavy traffic noise, can elicit emotional disorders in humans. However, the underlying neural substrate is still poorly understood. We exposed mice to moderate white noise for 28 days to induce anxiety-like behaviors, measured by open-field, elevated plus maze, and light-dark box tests. Invivo multi-electrode recordings in awake mice were used to examine neuronal activity. Chemogenetics were used to silence specific brain regions. Viral tracing, immunofluorescence, and confocal imaging were applied to define the neural circuit and characterize the morphology of microglia. Exposure to moderate noise for 28 days at an 85-dB sound pressure level resulted in anxiety-like behaviors in open-field, elevated plus maze, and light-dark box tests. Viral tracing revealed that fibers projecting from the auditory cortex and auditory thalamus terminate in the lateral amygdala (LA). A noise-induced increase in spontaneous firing rates of the LA and blockade of noise-evoked anxiety-like behaviors by chemogenetic inhibition of LA glutamatergic neurons together confirmed that the LA plays a critical role in noise-induced anxiety. Noise-exposed animals were more vulnerable to anxiety induced by acute noise stressors than control mice. In addition to these behavioral abnormalities, ionized calcium-binding adaptor molecule 1 (Iba-1)-positive microglia in the LA underwent corresponding morphological modifications, including reduced process length and branching and increased soma size following noise exposure. Treatment with minocycline to suppress microglia inhibited noise-associated changes in microglial morphology, neuronal electrophysiological activity, and behavioral changes. Furthermore, microglia-mediated synaptic phagocytosis favored inhibitory synapses, which can cause an imbalance between excitation and inhibition, leading to anxiety-like behaviors. Our study identifies LA microglial activation as a critical mediator of noise-induced anxiety-like behaviors, leading to neuronal and behavioral changes through selective synapse phagocytosis. Our results highlight the pivotal but previously unrecognized roles of LA microglia in chronic moderate noise-induced behavioral changes.

Clock knockout in inhibitory neurons reduces predisposition to epilepsy and influences anxiety-like behaviors in mice

Epilepsy is a brain disorder affecting up to 1 in 26 individuals. Despite its clinical importance, the molecular mechanisms of epileptogenesis are still far from clarified. Our previous study showed that disruption of Clock in excitatory neurons alters cortical circuits and leads to generation of focal epilepsy. In this study, a GAD-Cre;Clockflox/flox mouse line with conditional Clock gene knockout in inhibitory neurons was established. We observed that seizure latency was prolonged, the severity and mortality of pilocarpine-induced seizure were significantly reduced, and memory was improved in GAD-Cre;Clockflox/flox mice. We hypothesize that mice with CLOCK knockout in inhibitory neurons have increased threshold for seizure, opposite from mice with CLOCK knockout in excitatory neurons. Further investigation showed Clock knockout in inhibitory neurons upregulated the basal protein level of ARC, a synaptic plasticity-associated immediate-early gene product, likely through the BDNF-ERK pathway. Altered basal levels of ARC may play an important role in epileptogenesis after Clock deletion in inhibitory neurons. Although sEPSCs and intrinsic properties of layer 5 pyramidal neurons in the somatosensory cortex exhibit no changes, the spine density increased in apical dendrite of pyramidal neurons in CLOCK knockout group. Our results suggest an underlying mechanism by which the circadian protein CLOCK in inhibitory neurons participates in neuronal activity and regulates the predisposition to epilepsy.

Regulation of miRNA expression in the prefrontal cortex by fecal microbiota transplantation in anxiety-like mice.

The gut-brain axis and gut microbiota have emerged as key players in emotional disorders. Recent studies suggest that alterations in gut microbiota may impact psychiatric symptoms through brain miRNA along the gut-brain axis. However, direct evidence linking gut microbiota to the pathophysiology of generalized anxiety disorder (GAD) via brain miRNA is limited. In this study, we explored the effects of fecal microbiota transplantation (FMT) from GAD donors on gut microbiota and prefrontal cortex miRNA in recipient mice, aiming to understand the relationship between these two factors. Anxiety scores and gut microbiota composition were assessed in GAD patients, and their fecal samples were utilized for FMT in C57BL/6J mice. Anxiety-like behavior in mice was evaluated using open field and elevated plus maze tests. High-throughput sequencing of gut microbiota 16S rRNA and prefrontal cortex miRNA was performed. The fecal microbiota of GAD patients exhibited a distinct microbial structure compared to the healthy group, characterized by a significant decrease in Verrucomicrobia and Akkermansia, and a significant increase in Actinobacteria and Bacteroides. Subsequent FMT from GAD patients to mice induced anxiety-like behavior in recipients. Detailed analysis of gut microbiota composition revealed lower abundances of Verrucomicrobia, Akkermansia, Bifidobacterium, and Butyricimonas, and higher abundances of Deferribacteres, Allobaculum, Bacteroides, and Clostridium in mice that received FMT from GAD patients. MiRNA analysis identified five key miRNAs affecting GAD pathogenesis, including mmu-miR-10a-5p, mmu-miR-1224-5p, mmu-miR-218-5p, mmu-miR-10b-5p, and mmu-miR-488-3p. Notably, mmu-miR-488-3p showed a strong negative correlation with Verrucomicrobia and Akkermansia. This study demonstrates that anxiety-like behavior induced by human FMT can be transmitted through gut microbiota and is associated with miRNA expression in the prefrontal cortex. It is inferred that the reduction of Akkermansia caused by FMT from GAD patients leads to the upregulation of mmu-miR-488-3p expression, resulting in the downregulation of its downstream target gene Creb1 and interference with its related signaling pathway. These findings highlight the gut microbiota's crucial role in the GAD pathophysiology.

Post-Weaning Social Isolation Alters Puberty Onset by Suppressing Electrical Activity of Arcuate Kisspeptin Neurons.

Post-weaning social isolation (PWSI) in rodents is an advanced psychosocial stress model in early life. Some psychosocial stress, such as restrain and isolation, disrupt reproductive physiology in young and adult period. Mechanisms of early life stress (ELS) effects on central regulation of reproduction needs to be elucidated. We have investigated the effects of PWSI on function of Arcuate Kisspeptin (ARCKISS1) neurons by using electrophysiological techniques combining with monitoring of puberty onset and estrous cycle in male and female Kiss1-Cre mice. Female mice were monitored for puberty onset with vaginal opening examination during social isolation. After isolation, estrous cycle of female mice was monitored with vaginal cytology. Anxiety-like behavior of mice was determined by elevated plus maze test. Effects of PWSI on electrophysiology of ARCKISS1 neurons were investigated by patch clamp method after intracranial injection of AAV-GFP virus into arcuate nucleus of Kiss1-Cre mice after isolation period. We found that both male and female isolated mice showed anxiety-like behavior. PWSI caused delay in vaginal opening and extension in estrous cycle length. Spontaneous firing rate of ARCKISS1 neurons were significantly lower in the isolated male and female mice. The peak amplitude of inhibitory post-synaptic currents to ARCKISS1 neurons was higher in the isolated mice while frequency of excitatory post-synaptic currents was higher in group housed mice. These findings demonstrate that PWSI alters pre- and post- pubertal reproductive physiology through metabolic and electrophysiological pathways.

Chemotherapeutic drug elemene induces pain and anxiety-like behaviors by activating GABAergic neurons in the lateral septum of mice

Most chemotherapeutic drugs are potent and have a very narrow range of dose safety and efficacy, most of which can cause many side effects. Chemotherapy-induced peripheral neuropathy (CIPN) is the most common and serious side effect of chemotherapy for cancer treatment. However, its mechanism of action is yet to be fully elucidated. In the present study, we found that the treatment of the chemotherapy drug elemene induced hyperalgesia accompanied by anxiety-like emotions in mice based on several pain behavioral assays, such as mechanical allodynia and thermal hyperalgesia tests. Second, immunostaining for c-fos (a marker of activated neurons) further showed that elemene treatment activated several brain regions, including the lateral septum (LS), cingulate cortex (ACC), paraventricular nucleus of the thalamus (PVT), and dorsomedial hypothalamic nucleus (DMH), most notably in the GABAergic neurons of the lateral septum (LS). Finally, we found that both chemogenetic inhibition and apoptosis of LS neurons significantly reduced pain- and anxiety-like behaviors in mice treated with elemene. Taken together, these findings suggest that LS is involved in the regulation of elemene-induced chemotherapy pain and anxiety-like behaviors, providing a new target for the treatment of chemotherapy pain induced by elemene.

An ancient polymorphic regulatory region within the BDNF gene associated with obesity modulates anxiety-like behaviour in mice and humans.

Obesity and anxiety are morbidities notable for their increased impact on society during the recent COVID-19 pandemic. Understanding the mechanisms governing susceptibility to these conditions will increase our quality of life and resilience to future pandemics. In the current study, we explored the function of a highly conserved regulatory region (BE5.1) within the BDNF gene that harbours a polymorphism strongly associated with obesity (rs10767664; p = 4.69 × 10-26). Analysis in primary cells suggested that the major T-allele of BE5.1 was an enhancer, whereas the obesity-associated A-allele was not. However, CRISPR/CAS9 deletion of BE5.1 from the mouse genome (BE5.1KO) produced no significant effect on the expression of BDNF transcripts in the hypothalamus, no change in weight gain after 28 days and only a marginally significant increase in food intake. Nevertheless, transcripts were significantly increased in the amygdala of female mice and elevated zero maze and marble-burying tests demonstrated a significant increase in anxiety-like behaviour that could be reversed by diazepam. Consistent with these observations, human GWAS cohort analysis demonstrated a significant association between rs10767664 and anxiousness in human populations. Intriguingly, interrogation of the human GTEx eQTL database demonstrated no effect on BDNF mRNA levels associated with rs10767664 but a highly significant effect on BDNF-antisense (BDNF-AS) gene expression and splicing. The subsequent observation that deletion of BE5.1 also significantly reduced BDNF-AS expression in mice suggests a novel mechanism in the regulation of BDNF expression common to mice and humans, which contributes to the modulation of mood and anxiety in both species.

The delta opioid receptor agonist KNT-127 relieves innate anxiety-like behavior in mice by suppressing transmission from the prelimbic cortex to basolateral amygdala.

Excitatory projections from the prelimbic cortex (PL) to the basolateral nucleus of the amygdala (BLA) are implicated in the regulation of anxiety-like behaviors, and we previously demonstrated that anxiolytic-like effects of the selective delta-opioid receptor (DOP) agonist KNT-127 is involved in suppressing glutamate neurotransmission in the PL. Here, we investigated the mechanisms underlying the anxiolytic-like effect of KNT-127 in mice by combining optogenetic stimulation of the PL-BLA pathway with behavioral analyses. Four-week-old male C57BL/6J mice received bilateral administration of adeno-associated virus (AAV)2-CaMKIIa-hChR2(H134R)-enhanced yellow fluorescent protein (EYFP) into the PL to induce expression of the light-activated excitatory ionic channel ChR2. Subsequently, an optic fiber cannula connected to a wireless photo-stimulator was implanted into the BLA for optogenetic PL-BLA pathway stimulation. We evaluated innate anxiety using the elevated plus maze (EPM) and open field (OF) tests as well as learned anxiety using the contextual fear conditioning (CFC) test. Optogenetic activation of the PL-BLA pathway enhanced anxiety-like behaviors in the EPM and OF, while prior subcutaneous administration of KNT-127 (10 mg/kg) reduced this anxiogenic effect. In contrast, optogenetic activation of the PL-BLA pathway had no significant effect on conditioned fear. Our findings indicate that the PL-BLA circuit contributes to innate anxiety and that the anxiolytic-like effects of KNT-127 are mediated at least in part by suppression of PL-BLA transmission. The PL delta-opioid receptor may thus be an effective therapeutic target for anxiety disorders.

Timp1 Deletion Induces Anxiety-like Behavior in Mice.

The hippocampus is essential for learning and memory, but it also plays an important role in regulating emotional behavior, as hippocampal excitability and plasticity affect anxiety and fear. Brain synaptic plasticity may be regulated by tissue inhibitor of matrix metalloproteinase 1 (TIMP1), a known protein inhibitor of extracellular matrix (ECM), and the expression of TIMP1 in the hippocampus can be induced by neuronal excitation and various stimuli. However, the involvement of Timp1 in fear learning, anxiety, and hippocampal synaptic function remains to be established. Our study of Timp1 function in vivo revealed that Timp1 knockout mice exhibit anxiety-like behavior but normal fear learning. Electrophysiological results suggested that Timp1 knockout mice showed hyperactivity in the ventral CA1 region, but the basic synaptic transmission and plasticity were normal in the Schaffer collateral pathway. Taken together, our results suggest that deletion of Timp1 in vivo leads to the occurrence of anxiety behaviors, but that Timp1 is not crucial for fear learning.

Foxg1 Modulation of the Prkcd Gene in the Lateral Habenula Mediates Trigeminal Neuralgia-Associated Anxiety-Like Behaviors in Mice

Foxg1 Modulation of the Prkcd Gene in the Lateral Habenula Mediates Trigeminal Neuralgia-Associated Anxiety-Like Behaviors in Mice

Palmitoylated Sept8-204 modulates learning and anxiety by regulating filopodia arborization and actin dynamics.

Septin proteins are involved in diverse physiological functions, including the formation of specialized cytoskeletal structures. Septin 8 (Sept8) is implicated in spine morphogenesis and dendritic branching through palmitoylation. We explored the role and regulation of a Sept8 variant in human neural-like cells and in the mouse brain. We identified Sept8-204 as a brain-specific variant of Sept8 that was abundant in neurons and modified by palmitoylation, specifically at Cys469, Cys470, and Cys472. Sept8-204 palmitoylation was mediated by the palmitoyltransferase ZDHHC7 and was removed by the depalmitoylase PPT1. Palmitoylation of Sept8-204 bound to F-actin and induced cytoskeletal dynamics to promote the outgrowth of filopodia in N2a cells and the arborization of neurites in hippocampal neurons. In contrast, a Sept8-204 variant that could not be palmitoylated because of mutation of all three Cys residues (Sept8-204-3CA) lost its ability to bind F-actin, and expression of this mutant did not promote morphological changes. Genetic deletion of Sept8, Sept8-204, or Zdhhc7 caused deficits in learning and memory and promoted anxiety-like behaviors in mice. Our findings provide greater insight into the regulation of Sept8-204 by palmitoylation and its role in neuronal morphology and function in relation to cognition.

Projections from anteromedial thalamus nucleus to the midcingulate cortex mediate pain and anxiety-like behaviors in mice

Prior research has demonstrated the involvement of the midcingulate cortex (MCC) and its downstream pathway in pain regulation. However, the mechanism via which pain information is conveyed to the MCC remains unclear. The present study utilized immunohistochemistry, chemogenetics, optogenetics, and behavior detection methods to explore the involvement of MCC, anteromedial thalamus nucleus (AM), and AM-MCC pathway in pain and emotional regulation. Chemogenetics or optogenetics methods were employed to activate/inhibit MCCCaMKIIα, AMCaMKIIα, AMCaMKIIα-MCC pathway. This manipulation evokes/relieves mechanical and partial heat hyperalgesia, as well as anxiety-like behaviors. In the complete Freund,s adjuvant (CFA) inflammatory pain model, chemogenetic inhibition of the AMCaMKIIα-MCCCaMKIIα pathway contributed to pain relief. Notably, this study presented the first evidence implicating the AM in the regulation of nociception and negative emotions. Additionally, it was observed that the MCC primarily receives projections from the AM, highlighting the crucial role of this pathway in the transmission of pain and emotional information.

The Analgesia Effect of Aucubin on CFA-Induced Inflammatory Pain by Inhibiting Glial Cells Activation-Mediated Inflammatory Response via Activating Mitophagy.

Inflammatory pain, characterized by sustained nociceptive hypersensitivity, represents one of the most prevalent conditions in both daily life and clinical settings. Aucubin, a natural plant iridoid glycoside, possesses potent biological effects, encompassing anti-inflammatory, antioxidant, and neuroprotective properties. However, its impact on inflammatory pain remains unclear. The aim of this study is to investigate the therapeutic effects and underlying mechanism of aucubin in addressing inflammatory pain induced by complete Freund's adjuvant (CFA). The CFA-induced inflammatory pain model was employed to assess whether aucubin exerts analgesic effects and its potential mechanisms. Behavioral tests evaluated mechanical and thermal hyperalgesia as well as anxiety-like behaviors in mice. The activation of spinal glial cells and the expression of pro-inflammatory cytokines were examined to evaluate neuroinflammation. Additionally, RNA sequencing was utilized for the identification of differentially expressed genes (DEGs). Molecular biology experiments were conducted to determine the levels of the PINK1 gene and autophagy-related genes, along with PINK1 distribution in neural cells. Furthermore, mitophagy induced by carbonyl cyanide m-chlorophenylhydrazone (CCCP) was employed to examine the roles of PINK1 and mitophagy in pain processing. Aucubin significantly ameliorated pain and anxiety-like behaviors induced by CFA in mice and reduced spinal inflammation. RNA sequencing indicated PINK1 as a pivotal gene, and aucubin treatment led to a significant downregulation of PINK1 expression. Further GO and KEGG analyses suggested the involvement of mitochondrial function in the therapeutic regulation of aucubin. Western blotting revealed that aucubin markedly decreased PINK1, Parkin, and p62 levels while increasing LC3B expression. Immunofluorescence showed the predominant co-localization of PINK1 with neuronal cells. Moreover, CCCP-induced mitophagy alleviated mechanical and thermal hyperalgesia caused by CFA and reversed CFA-induced mitochondrial dysfunction. In summary, our data suggest that aucubin effectively alleviates CFA-induced inflammatory pain, potentially through triggering the PINK1 pathway, promoting mitophagy, and suppressing inflammation. These results provide a novel theoretical foundation for addressing the treatment of inflammatory pain.

Clavulanic Acid Improves Memory Dysfunction and Anxiety Behaviors through Upregulating Glutamatergic Transporters in the Nucleus Accumbens of Mice Repeatedly Exposed to Khat Extract.

Khat (Catha edulis) is an evergreen shrub whose buds and leaves give a state of delight and euphoria when chewed. Cathinone, an amphetamine-like stimulant that is among the active ingredients in khat, is able to downregulate glutamate transporter subtype I (GLT-1). Neurobehavioral dysfunctions such as altered locomotor activity, anorexia, and nociception have been observed in animals exposed to cathinone. Interestingly, treatment with a β-lactam antibiotic such as ceftriaxone, which upregulates GLT-1, normalizes cathinone-induced conditioned place preference, and alters repetitive movements in rats. However, little is known about the role of the glutamatergic system in memory dysfunction and anxiety-like behaviors in mice exposed to khat. We found here that clavulanic acid, a β-lactam-containing compound and GLT-1 upregulator, would modulate the neurobehavioral changes, including memory impairment and anxiety-like behaviors, associated with repeated exposure of mice to khat. Our data supported that clavulanic acid could improve memory impairment and anxiety-like behaviors through upregulating GLT-1 in the nucleus accumbens (NAc), an effect abolished with a selective GLT-1 blocker. This upregulation was associated with restored glutamate/cystine antiporter expression in the NAc using a Western blotting assay. Cathine and cathinone were identified in khat extract using the gas chromatography technique. Our work provides preclinical insight into the efficacy of β-lactam-containing compounds for the attenuation of neurobehavioral changes induced by khat exposure.