General anesthetics can exert significant adverse effects on the central nervous system. This study aimed to investigate whether repeated exposure to sevoflurane induces depression-like behaviors in postpartum rats. Pregnant rats were exposed to 3% sevoflurane for 2 h on gestational days 13-15. Emotional behaviors were assessed on postpartum days 1, 7, 14, and 21. Hippocampal protein levels associated with the AMPK/SIRT1/NLRP3 signaling pathway were analyzed by Western blotting. Microglial activation and inflammasome expression were analyzed by immunofluorescence, and cytokine levels (IL-1β, IL-18, TNF-α) by ELISA. To explore the role of the AMPK/SIRT1/NLRP3 pathway and neuroinflammation in postpartum maternal depression, rats were treated with AICAR (an AMPK agonist), MCC950 (an NLRP3 antagonist), and minocycline (a microglial activation inhibitor). Additionally, ketamine, with or without dorsomorphin (an AMPK antagonist), was administered to assess whether ketamine's antidepressant effects are mediated through this pathway. Sevoflurane-exposed rats exhibited behavioral impairments on postpartum day 1, including increased immobility in the forced swim test, prolonged feeding latency, reduced food consumption in the novelty-suppressed feeding test, and decreased movement in the open field test. These behaviors were accompanied by decreased AMPK/SIRT1 expression, NLRP3 inflammasome activation, and microglial activation in the hippocampus, resulting in significant inflammatory cytokine release. Treatment with AICAR, MCC950, minocycline, or ketamine alleviated these effects, while dorsomorphin reversed the antidepressant effects of ketamine. Our findings indicate that repeated sevoflurane exposure during mid-gestation induces depression-like behaviors in postpartum rats, and that ketamine alleviates these behaviors by reducing microglial neuroinflammation and NLRP3 inflammasome activation via the AMPK/SIRT1 signaling pathway.

The recent global spread of monkeypox virus (MPXV) highlights the urgent need for effective antiviral therapies. Through high-throughput screening of an FDA-approved small molecule drug library, we identified salinomycin as a potent inhibitor of orthopoxvirus infection. Salinomycin showed nanomolar anti-MPXV activity in human keratinocytes (HaCaT) and lung epithelial cells (A549), with selectivity indices >100. Importantly, its efficacy was validated in primary human fibroblasts, reducing infectious viral titers by ∼2.6 log units (440-fold) versus controls. Mechanistic studies revealed salinomycin acts at the viral post-entry stage, blocking membrane fusion via disrupting endosomal acidification, without affecting clathrin-mediated endocytosis. This host-directed mechanism was validated by DiO fusion assays, acridine orange staining, and low-pH pulse rescue experiments. In a lethal intranasal vaccinia virus (VACV-WR) murine challenge model, oral salinomycin (1 mg/kg/day) significantly reduced lung viral loads, mitigated histopathology, and conferred full mortality protection. Furthermore, salinomycin synergized with tecovirimat, an approved viral egress inhibitor. In fixed-ratio combinations, salinomycin's EC50 dropped ∼4.0-fold (30.12 nM to 7.50 nM), and tecovirimat's ∼3.2-fold (11.96 nM to 3.73 nM). Bliss independence analysis confirmed strong synergy, with a maximum ΔBliss value of 16.97. Notably, the combination achieved 100% survival in a high-dose VACV-WR model, an effect unmatched by either monotherapy. Collectively, these preliminary in vitro and in vivo data identify salinomycin as a potential host-targeted anti-orthopoxvirus inhibitor, especially in combination with tecovirimat. Further comprehensive preclinical evaluations, including full toxicological profiling and validation in clinically relevant models, are needed to define its translational potential for MPXV and other orthopoxvirus infections.

General anesthesia exposure in early life may disrupt the normal progression of developmental myelination, but the underlying mechanisms remain unclear. Early postnatal microglia in developing white matter exhibit diverse transcriptional and functional states, including a population with pronounced phagocytic activity. This study aims to investigate whether sevoflurane impairs oligodendrocyte myelination by promoting microglial phagocytosis of oligodendrocyte precursor cells (OPCs). Mice received either a single 2-h exposure to 3.3

ABSTRACT Dengue virus (DENV) infection poses a significant global health threat, and current prevention and treatment strategies are limited by challenges of lacking effective mosquito control measures and antibody‐dependent enhancement. This study reports that bacterial extracellular vesicles (BEVs) secreted by a soil bacterium Chryseobacterium aquifrigidense M24 exhibit potent anti‐DENV activity by triggering the structural disintegration of DENV particles prior to cellular entry in a dose‐dependent manner. Mechanistic investigations revealed that BEVs interact with the viral envelope, inducing premature membrane fusion. This process is characterized by reduced membrane fluidity and irreversible lipid rearrangement, leading to a significant increase in particle density, as shown by iodixanol gradient ultracentrifugation. The proposed ‘fusion‐triggered structural disruption’ is further supported by the induction of aberrant E protein oligomerization and morphological changes observed via transmission electron microscopy. This mechanism is specific to enveloped viruses, as BEVs showed no effect on non‐enveloped Enterovirus 71. Crucially, this BEV‐mediated inactivation extends to other enveloped viruses, including HCV, WNV and YFV, indicating broad‐spectrum potential. Our findings reveal a previously unexplored function of BEVs as virucidal agents, proposing a new ‘virus‐destructor’ strategy that contrasts with conventional fusion inhibitors and offering promising avenues for developing broad‐spectrum antiviral drugs.

RNA viruses, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), flaviviruses, and alphaviruses, represent a major source of emerging human infectious diseases. They pose a persistent threat to public health; however, few therapeutic options are available for severe infections. Through a natural product screening campaign, we identified ansatrienin B as a broad-spectrum inhibitor of multiple RNA viruses, including SARS-CoV-2, flaviviruses (e.g., YFV, WNV, DENV), and alphaviruses (e.g., CHIKV). Time-of-drug-addition assays indicated that ansatrienin B acts at both the early (entry) and intermediate (replication) stages of the viral life cycle. Surface plasmon resonance (SPR) and molecular docking studies validated a direct interaction between ansatrienin B and the RNA-dependent RNA polymerase (RdRp) of SARS-CoV-2 and WNV. Combined RNA pull-down and RdRp enzymatic activity assays (in gel, solution, and cellular forms) further demonstrated that ansatrienin B disrupts both the binding of RdRp to viral RNA and its enzymatic activity. In vivo, ansatrienin B showed significant efficacy in mouse models infected with SARS-CoV-2 or WNV infection. To facilitate screening and elucidate the structure-activity relationship (SAR), we generated a focused ansatrienin library via a mutasynthetic approach. Supplementation of four 3,5-AHBA analogs into a ΔmycB1-B4 mutant strain of Streptomyces flaveolus yielded 30 novel ansatrienin derivatives. Evaluation of anti-SARS-CoV-2 activity identified four analogs with enhanced potency, enabling the establishment of a preliminary SAR. Collectively, these findings establish ansatrienin B as a novel inhibitor targeting RdRp and provide a foundation for the development alternative broad-spectrum antiviral agents.

Chikungunya virus (CHIKV), a mosquito-borne alphavirus, causes debilitating febrile and arthritic disease and remains a persistent public health threat in tropical and subtropical regions, with no clinically approved antiviral drugs currently available, which underscores the urgent need for targeted and effective therapeutic interventions. Through high-throughput screening of an FDA-approved compound library, we identified retinoic acid (RA) as a broad-spectrum inhibitor of multiple arboviruses, exhibiting potent activity against CHIKV. Time-of-addition experiments, together with assays on viral binding, endocytosis, membrane fusion, replication and translation, were performed to determine the specific lifecycle stages inhibited by RA. Notably, RA exerts anti-CHIKV effects by selectively targeting eukaryotic translation initiation factor 4B (EIF4B), thereby disrupting the viral translation, as revealed by limited proteolysis-mass spectrometry (LiP-MS). And, our results demonstrated that RA administration exerted potent protective effects against CHIKV infection in vivo. Specifically, RA significantly reduced cerebral pathological damage, relieved clinical manifestations, and enhanced survival in a murine model of CHIKV-induced encephalitis, while also markedly attenuating footpad swelling and joint pathological alterations in a CHIKV-induced arthritis mouse model. Collectively, our findings highlight RA as a promising anti-CHIKV candidate targeting EIF4B, supporting its further development as a therapeutic agent against CHIKV infection.

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that causes acute febrile illness and chronic arthralgia, yet no approved antivirals exist. Although MXRA8 was previously identified as a CHIKV receptor, it cannot account for viral infection in MXRA8 absence both in vitro and in vivo , suggesting additional entry mediators. Here, we identify neuropilin-2 (NRP2) as a functional entry receptor for CHIKV. Through targeted siRNA screening in neural cells, we found NRP2 to be critical for viral entry independently of MXRA8. The extracellular domain of NRP2 binds directly to the CHIKV E2 glycoprotein, with defined amino acid residues mediating this interaction. NRP2 and MXRA8 function additively and non-competitively, expanding the known cellular entry landscape for CHIKV. Monoclonal antibodies targeting NRP2 or a soluble NRP2-Fc decoy potently inhibit CHIKV infection in vitro and in vivo , moreover attenuate joint pathology in a murine model. These findings establish NRP2 as a key entry receptor for CHIKV, explaining its broad tissue tropism and presenting a promising target for therapeutic intervention.

BACKGROUND AND PURPOSE:Chikungunya virus (CHIKV) causes severe acute and chronic disease, yet no approved specific antiviral treatment exists. To rapidly identify potential treatments, we aimed to screen an FDA-approved drug library for inhibitors of CHIKV infection. Tirbanibulin, a dual-microtubule polymerization and Src kinase inhibitor, was assessed for its potential anti-CHIKV activity. METHODS:We performed a high-throughput screen of an FDA-approved drug library. Mechanism-of-action studies included entry-step analysis, surface plasmon resonance (SPR) binding assays and molecular docking. In vivo efficacy was evaluated in lethal murine neuroinfection and CHIKV-induced arthritis models following oral administration of the candidate compound. EXPERIMENTAL APPROACH:Tirbanibulin exhibited nanomolar to low-micromolar antiviral activity (EC50 range: 0.035-75.64 μM) across multiple cell lines, with high selective indices in key target HT22 and Huh7 cells. It acted at a post-attachment entry step, inhibiting clathrin-mediated endocytosis and potentially viral fusion. Surface plasmon resonance confirmed direct, high-affinity binding to the CHIKV E1 and E2 glycoprotein complex (K_D = 73 nM). In the lethal neuroinfection model, oral tirbanibulin significantly improved survival and reduced brain viral loads; in the arthritis model, it markedly attenuated footpad swelling and inflammatory pathology. CONCLUSION AND IMPLICATIONS:Tirbanibulin is a novel, orally bioavailable entry-stage inhibitor that directly targets the E2 glycoprotein. Multiple preclinical and clinical studies have confirmed its favourable oral bioavailability and safety. Given its established clinical safety profile, it represents a promising repurposing candidate for clinical evaluation against Chikungunya fever.

Human rhinoviruses (HRVs) are major etiological agents of upper respiratory tract infections and are strongly associated with asthma exacerbations and chronic obstructive pulmonary disease, yet no approved antiviral therapies are currently available. Reverse genetics systems, particularly reporter viruses, are critical for facilitating antiviral discovery; however, robust and broadly applicable HRV infectious clones remain limited. Here, we recovered a viral genome from a clinical sample and generated infectious cDNA clones of an HRV-A1 strain, designated HRV-fd. Both the wild-type virus and a Nanoluc (Nluc) reporter virus exhibited efficient replication in HeLa cells without inducing pronounced cytopathic effects and were capable of infecting HeLa-ICAM1 cells at extremely low inocula. Leveraging the reporter virus, we conducted a drug-repurposing screen and identified pyrvinium pamoate as an antiviral compound that targets multiple steps of the HRV infection cycle, exhibiting an IC50 of 58.6 nM, an IC90 of 193.1 nM, and a CC50 of 3.4 μM. Notably, pyrvinium pamoate conferred sustained antiviral activity following transient exposure and subsequent withdrawal, suggesting the induction of host cellular alterations that restrict viral amplification. Collectively, these findings establish a robust HRV-A1 reverse genetics and reporter platform for virological studies and antiviral screening, and identify pyvinium pamoate as a promising lead compound for the development of effective anti-HRV therapeutics.

Purpose:This study aims to investigate the factors influencing the impact of healthcare team collaboration on vaccination status among patients with Chronic Obstructive Pulmonary Disease (COPD) in primary care settings in Beijing, China. Patients and Methods:A multicenter cross-sectional survey was conducted among members of family doctor teams in Beijing from October to December 2025. A structured questionnaire assessed team collaboration behaviors, influencing factors, guideline awareness, and vaccination practices. Descriptive statistics and chi-square tests were performed. Results:A total of 209 valid responses were included (response rate: 92.07%). Although 57.9% of participants supported shared responsibility for vaccination, only 24.9% reported frequent team discussions, and 9.1% reported none in the past three months. Unstable vaccine supply (M=2.83) and limited performance incentives (M=3.76) were key barriers. While 66.5% of respondents were familiar with relevant guidelines, only 44.5% consistently provided strong vaccination recommendations in clinical scenarios. Patient hesitancy was primarily attributed to concerns about side effects (69.4%). Most respondents (67.0%) estimated vaccination coverage among COPD patients to be below 30%. Conclusion:A substantial gap exists between guideline awareness and implementation of vaccination in primary care COPD management. Limited team communication, inadequate system support, and patient hesitancy jointly constrain vaccination uptake. Strengthening structured team collaboration, improving incentive mechanisms, and enhancing scenario-based guideline training may help improve vaccination coverage in this population.

Effective respiratory mucosal vaccines remain urgently needed to mitigate the rapid mutation and transmission of SARS-CoV-2. Here, we demonstrated that the spike protein (S-2P) of ancestral SARS-CoV-2 acted as a self-adjuvanted antigen for intranasal immunization, inducing robust systemic and mucosal immunity via integrin- and STING-dependent pathways. In contrast, H1N1 influenza hemagglutinin (HA) failed to generate measurable serum IgG or mucosal IgA following intranasal immunization. In mice, intranasal S-2P vaccination conferred complete protection against lethal ancestral SARS-CoV-2 challenge and partial cross-protection against heterologous Omicron variants, with both effects being IFN-γ- and CD8 + T cell-dependent. Co-administration of S-2P with the clinical immunomodulator lentinan (LNT) achieved complete protection against Omicron variants, mediated by IFN-γ but largely independent of CD8 + T cells. These findings establish S-2P + LNT as a safe, broad-spectrum mucosal vaccine candidate against emerging SARS-CoV-2 variants and reveal novel protection mechanisms beyond neutralizing antibodies and T cell immunity.

Women with term pregnancies undergoing cesarean delivery under combined spinal and epidural anesthesia (CSEA) usually present with hemodynamic variability and a high incidence of hypotension. Among the commonly used anesthetic drugs for CSEA, ropivacaine reduces the incidence of hypotension compared with bupivacaine. However, most studies have focused on blood pressure and heart rate measurements to evaluate the effects of these two anesthetic drugs on parturients. This study aimed to compare maternal cardiac output and hemodynamic parameters, such as cardiac preload and afterload, of ropivacaine and bupivacaine using transthoracic echocardiography. Eighty-two parturients scheduled for elective caesarean deliveries under CSEA anesthesia were randomly assigned to receive 0.5

Climate change has expanded the range of vectors for the chikungunya virus (CHIKV), thereby increasing its global threat. This situation underscores the urgent need for antiviral therapies. However, the development of drugs is limited by the need for Biosafety Level 3 (BSL-3) containment and the lack of tools for real-time infection monitoring. To address these challenges, we developed a replication-competent reporter virus called CHIKV-ΔnsP3-Nluc. We inserted the Nano-luciferase (Nluc) gene into the attenuation site of the FDA-approved live-attenuated vaccine strain VLA1553 (IXCHIQ). This modified virus retains an attenuated phenotype, replicating more slowly than wild-type CHIKV while maintaining high genetic stability. Nluc activity correlates well with viral titer (R2 = 0.9689), enabling precise quantification of viral replication. In interferon-deficient A6 mice, CHIKV-ΔnsP3-Nluc caused a lethal yet attenuated infection. Real-time bioluminescence imaging revealed that the spleen and kidneys were the primary replication sites, with luminescence intensity closely matching viral titers (R2 = 0.8388). We confirmed the platform's potential for rapid antiviral screening using the nucleoside analog 4'-fluorouridine (4'-FlU). This compound inhibited viral replication in vitro (EC50= 0.063 µM) and significantly reduced viremia and mortality in vivo at a dose of 10 mg/kg, resulting in 100% survival. CHIKV-ΔnsP3-Nluc is a BSL-2-compliant tool that combines the safety of a vaccine candidate with a sensitive reporter function. It has the potential to accelerate studies of CHIKV pathogenesis and assist in developing new antiviral therapies.IMPORTANCEThe expanding range of CHIKV outbreaks poses a threat to global public health, and there is currently a lack of suitable tools for the development of antiviral drugs. In this study, we developed a Nano-luciferase reporter virus based on the attenuated vaccine strain VLA1553 to track viral infection. This viral assay can effectively evaluate the inhibitory and protective effects of candidate compounds both in vitro and in vivo, and its luciferase signal reliably reflects the level of viral replication. Our research provides a safe and stable tool that serves as a suitable platform for the development of antiviral drugs targeting CHIKV.

Background Perioperative benzodiazepines increase the risk of sensory hypersensitivity and agitation during recovery, but the neural mechanisms remain unclear. Auditory gating filters redundant information to prevent network overload. We hypothesise that benzodiazepines impair auditory gating during recovery, leading to heightened auditory responsiveness. Methods Simultaneous electroencephalography, electromyography, and behavioural analyses assessed the effects of remimazolam, a novel benzodiazepine, on arousal states. Multi-region microelectrodes and Neuropixels probes recorded cortical and subcortical local field potentials and single-unit activity. Optogenetics was applied to test the impact of remimazolam on auditory gating. Results Both spontaneous and paired-tone evoked neuronal activity were suppressed during anaesthesia and sedation, followed by rebound enhancement and auditory gating deficits during recovery (T2/T1 ratio in posterior parietal cortex: baseline 0.38 [0.01] vs recovery 0.82 [0.02], P<0.001; dorsal hippocampus: 0.34 [0.01] vs 1.10 [0.03], P<0.001; and mediodorsal thalamic nucleus: 0.48 [0.01] vs 1.12 [0.02], P<0.001]). Optogenetic manipulations of the prefrontal cortex and brainstem auditory nuclei revealed that both top-down and bottom-up inputs were blocked during anaesthesia. During recovery, top-down inputs normalised, whereas bottom-up inputs exceeded baseline, along with gating deficits. A subset of gamma-aminobutyric acid (GABA)ergic neurones in thalamic reticular nucleus exhibited sustained responses to paired tones. During recovery, both their proportion (63.0% vs 29.3%) and firing strength (maximum firing rate: 28.9 [3.9] vs 17.4 [3.0], P<0.001) were diminished, resulting in insufficient inhibition of bottom-up inputs and exaggerated responses to external stimuli. Conclusions Our findings elucidate the dynamic changes in sensory processing and the underlying mechanisms during benzodiazepine-induced anaesthesia and recovery, providing valuable insights for optimising clinical anaesthesia management and postoperative recovery strategies.

Background Our previous data suggested that autophagy is crucial for neuropathic pain. The different cell types and varying degrees of regulation of STING may lead to a paradoxical effect on pain and emotions in the neuropathic pain model. Up to now, whether STING modulates neuropathic pain in PrL neurons via ER-phagy is still unknown.Method In this study, we investigated the effect of ER-phagy in the prefrontal cortex (PrL) on neuropathic pain. We administered 4-phenylbutyric acid, tunicamycin, 3-methyladenine, and rapamycin to evaluate the interaction between endoplasmic reticulum (ER) stress and autophagy in the PrL of SNL (spinal nerve ligation). We injected AAV to investigate whether ER-phagy modulated pain and emotional behaviors. We further explored whether STING and its pathway as a modulation target for ER-phagy to participate in the pain process. We injected 2 ' 3-cGAMP and RU521 to modulate the cGAS/STING pathway in ER-phagy in SNL mice. Moreover, we modulated STING expression and regulated the levels of ER-phagy and the interaction between STING and LC3 in neurons through PrL AAV injections.Results The data indicated that ER-phagy alleviated the excessive ER stress induced by SNL in PrL through the cGAS/STING pathway. Regulating ER-phagy in PrL neurons through AAV tools altered pain and emotion-related behaviors. In addition, regulating STING in PrL neurons altered the comorbidity of pain and emotion. Importantly, the binding interaction between STING and LC3 in PrL neurons provides a novel target for PrL ER-phagy.Conclusion Enhanced ER-phagy of PrL neurons provides analgesic, anti-anxiety, and antidepressant effects through modulating STING in SNL mice.

IntroductionWest Nile virus (WNV)-associated neurological diseases pose a global public health burden, yet no approved antiviral treatments are available. This is primarily due to the challenges of crossing the blood-brain barrier and the lengthy, costly process of drug development.MethodsHigh-throughput screening of a blood-brain barrier-penetrating FDA-approved compound library was performed to identify anti-WNV candidates in a neuronal cell infection model. Antiviral efficacy was further evaluated across different target cells and a range of flaviviruses, as well as in a mouse model of central nervous system (CNS) WNV infection. Stage-of-action analysis and mechanistic studies were conducted.ResultsMiconazole emerged as a promising candidate with significant antiviral efficacy against multiple flaviviruses in different cell types. In the CNS infection mouse model, miconazole treatment reduced viral load in brain tissue and improved survival rates. The compound primarily inhibited viral replication, without affecting binding, internalization, or membrane fusion. Mechanistic studies suggested that this antiviral effect may be mediated through the downregulation of host protein thioredoxin-like 1 (TXNL1).DiscussionOur findings highlight miconazole as a promising candidate for repurposing in anti-flavivirus therapy and identify TXNL1 as a potential host target for the development of broad-spectrum antivirals.

Aflatoxin B1 (AFB1) contamination in edible oils poses a serious threat to food safety, yet its sensitive and rapid detection is hindered by the high viscosity and electrical insulativity of oil matrices. Herein, we constructed an electrochemiluminescence (ECL) sensing chip for rapid determination of AFB1 in corn oil. Honeycomb-like porous carbon materials (HPCMs) derived from wood waste were employed to encapsulate CsPbBr3 quantum dots (CsPbBr3 QDs). The resulting CsPbBr3 QDs@HPCMs composites exhibit efficient ECL responses without coreactant participation in 1-butyl-3-methylimidazolium hexafluorophosphate ionic liquid electrolyte. A patterned three-electrode chip fabricated with carbon ink on a plastic substrate was constructed as a portable ECL device. Coupled with a humic acid-bonded silica solid-phase extraction needle, this platform enables ultrasensitive and selective detection of AFB1 in corn oil. This study offers a promising ECL sensing strategy in oil-phase food safety analysis.

The SARS-CoV-2 spike (S) protein, a trimeric structure comprising three receptor binding domains (RBDs) and three N-terminal domains (NTDs), undergoes substantial conformational changes to a fusion-prone open state for angiotensin-converting enzyme 2 (ACE2) binding and host cell infection. Stabilizing its closed state is a key antiviral strategy but remains challenging. Here, we introduce S416, a novel amphipathic molecule acting as a "molecular bolt". Cryo-EM study reveals that S416 binds concurrently to six sites across two distinct druggable interfaces: three molecules at the RBD-RBD interfaces and three at the NTD-RBD interfaces. This unique "dual-locking" mechanism, driven by S416's polar carboxyl head and nonpolar phenylthiazole tail, robustly stabilizes the spike trimer in a locked, closed conformation through strong inter-domain interactions, reducing structural flexibility and atomic fluctuations compared to the apo structure resolved synchronously. Crucially, these RBD-RBD and NTD-RBD interfaces are conserved across human-infecting coronaviruses, suggesting potential as broad-spectrum antiviral targets. Our findings demonstrate that the highly dynamic spike trimer can be effectively stabilized by an amphipathic molecular bolt targeting both the inter- and intra-monomer interfaces, offering a promising strategy against emerging coronaviruses.

BACKGROUND:Noradrenergic projections from the locus coeruleus (LC) to the thalamus and anterior cingulate cortex (ACC) contribute to pain-like behaviors, yet their hierarchical organization remains unclear. This article examines how locus coeruleus-derived norepinephrine inputs to the paraventricular thalamic nucleus (PVA) and ACC differentially regulate nociceptive sensitization. METHODS:In adult male and female mice, complete Freund's adjuvant was used to induce pain-like behaviors. To examine functional connectivity among locus coeruleus, PVA, and ACC, targeted recombination in active populations (Fos-TRAP), in vivo recordings, and viral tracing were combined. Then optogenetic and chemogenetic tools were used to selectively manipulate locus coeruleus projections and assess their impact on neural activity and pain behaviors. RESULTS:Complete Freund's adjuvant led to enhanced c-Fos expression in locus coeruleus, PVA, and ACC (cells per microscopic field; locus coeruleus: 13.60 ± 2.24 vs. 44.50 ± 7.72; PVA: 8.00 ± 1.58 vs. 66.40 ± 9.45; ACC: 12.80 ± 2.28 vs. 36.70 ± 2.59; P < 0.001), alongside increased γ-band activity and single-unit firing rates. Monosynaptic LC-ACC and polysynaptic LC-PVA-ACC circuits were identified. Notably, nociception-related locus coeruleus neurons preferentially projected to PVA, which subsequently targeted hyperactive ACC neurons. Under inflammatory pain conditions, activation of the LC-PVA-ACC circuits evoked greater ACC firing (Hz; LC-PVA-ACC vs. LC-ACC: 15.75 ± 2.88 vs. 9.72 ± 2.06; P < 0.001) and tactile stimulus-evoked responses (Hz; 22.98 ± 2.60 vs. 15.34 ± 1.86; P < 0.001) than direct LC-ACC activation. Consistently, optogenetic or chemogenetic manipulation of the LC-PVA-ACC circuit produced stronger modulation of mechanical and thermal pain sensitivity than direct LC-ACC stimulation. CONCLUSIONS:This study identified the LC-PVA-ACC pathway as a hierarchical noradrenergic circuit that modulates nociceptive sensitization via a thalamocortical relay, thereby revealing a circuit-specific mechanism by which the locus coeruleus-norepinephrine system regulates pain processing.

The Omicron BA.2.86 subvariants, JN.1, KP.2, and KP.3, have become predominant globally, raising concerns about their immune evasion from vaccines and monoclonal antibody (mAb) treatments. These variants harbor more receptor-binding domain (RBD) mutations than the XBB and EG.5 sub-lineages, which are already known to compromise vaccine and therapeutic efficacy. We evaluated sera from individuals vaccinated with inactivated vaccines, with or without breakthrough infections, as well as COVID-19 convalescents. Our results showed a substantial decrease in serum neutralizing activity against the JN.1, KP.2, XBB.1.5, and EG.5.1 variants compared to BA.2. Additionally, we developed 19 neutralizing antibodies from memory B cells, with some retaining efficacy against earlier Omicron variants. However, potency was notably diminished against newer subvariants like BF.7, BQ.1, XBB.1.5, and BA.2.86. Of mAbs, those isolated from COVID-19 convalescents, particularly SA-3, exhibited exceptional potency across ten variants from BA.2 to KP.2, with IC50 values ranging from 0.006 to 2.546 μg/mL. However, SA-3 had lost neutralizing activity against the KP.3 due to the Q493E mutation, but the KP.3 became susceptible to neutralization by the other mAb, SA-6. In contrast, SA-6 was unable to neutralize KP.2 because of the presence of R346T mutation. Our findings underscore the importance of continuous surveillance of viral evolution and the need for updated vaccines and therapeutics to combat the ongoing evolution of SARS-CoV-2, particularly in the context of emerging variants that escape both vaccine-induced immunity and monoclonal antibody treatments.