Stress-related experiences alter glutamatergic signaling and neuronal excitability, but the mechanisms that couple experience to dentate granule cell function remain incompletely understood. Here, we examined how protein kinase N1a (PKN1a), a protein kinase C-like serine/threonine kinase, and repeated swim exposure regulate mouse hippocampal dentate granule cell excitability, with a focus on the neuronal glutamate transporter excitatory amino acid transporter 3 (EAAT3) and group I metabotropic glutamate receptors (mGluRs). Five days of repeated swim exposure increased spike firing in mature dentate granule cells from wild-type mice. PKN1a knockout produced a similar increase, and repeated swim did not further enhance firing in knockout mice. The enhanced firing observed after repeated swim exposure and in PKN1a knockout mice was reduced by co-application of an mGluR1 antagonist (LY367385) and an mGluR5 antagonist (MPEP). Inhibition of glutamate transporters with DL-TBOA increased granule cell firing in control wild-type mice but did not further increase firing in repeated-swim wild-type or PKN1a knockout mice, suggesting occlusion of transporter-dependent regulation of excitability. Repeated swim exposure and PKN1a knockout also reduced total and surface expression of EAAT3 in the hippocampus, whereas expression of the glial glutamate transporter EAAT2 was not significantly altered. Finally, PKN1a knockout and repeated swim exposure reduced anxiety-related behavior in the elevated plus maze test. Thus, PKN1a-dependent regulation of EAAT3 may restrain group I mGluR-dependent excitability in dentate granule cells, whereas repeated swim exposure and PKN1a knockout shift this system toward a lower-EAAT3, higher-excitability state accompanied by reduced anxiety-related behavior.

Oxycodone is an opioid analgesic metabolized by the CYP2D enzyme to oxymorphone. We investigated in rats the impact of CYP2D metabolism in the brain on oxycodone-induced reward and aversion, assessed by conditioned place preference (CPP). During conditioning rats were confined to one compartment for 60 minutes after gavage with oxycodone (or oxymorphone) or with water; treatment and compartments were alternated daily for six days. Rats then underwent 60 minutes of posttests for five days where CPP was assessed. Pretreatments with the CYP2D inhibitor propranolol, or vehicle, injected intracerebroventricularly 24 hours - prior to first and third of six conditioning sessions blunted CPP and with a trend for increased aversion to the oxycodone paired compartment. Pretreatment with the brain CYP2D enzyme inducer nicotine, for 12 days prior to oxycodone conditioning, did not change CPP to oxycodone but tended to reduced aversion to oxycodone paired compartment. Oral oxymorphone induced a CPP with no observed aversion; it was unaffected by CYP2D inhibitor pretreatment. Together this indicates that oral oxycodone and oral oxymorphone both induce CPP. Inhibiting brain CYP2D, previously shown to reduce the relative concentrations of oxycodone to oxymorphone, reduces CPP and tended to increase aversion suggesting that variation in CYP2D activity in the brain could be a source of interindividual differences in oxycodone induced reward and abuse liability.

The rising global burden of depression necessitates novel mechanistic insights and therapeutic targets. Voltage-gated sodium channel β2-subunit (Navβ2, coded by SCN2B) has been implicated in neuropsychiatric disorders, yet its role in depression remains unexplored. Here, we identify SCN2B as a previously unrecognized driver of depression-like pathophysiology. Using SCN2B-overexpressing (SCN2B-OV) mice across different ages (1, 12, and 20 months) and CRISPR- engineered Navβ2[V173A,V175K] mice with selective reduction of the Navβ2 intracellular domain (NICD), we combined behavioral phenotyping, electrophysiology, neurotransmitter profiling, and molecular analyses, with chronic unpredictable mild stress (CUMS) as an environmental challenge. SCN2B-OV mice exhibited accelerated depression-like phenotypes, including anhedonia (sucrose preference 52.42% reduction, P<0.05), behavioral despair (forced swim test immobility 67.42% increase, P<0.05), and cognitive deficits (Y-maze alternation 50.90% reduction, P<0.05), independent of natural aging. Electrophysiologically, SCN2B overexpression induced a striking prefrontal-hippocampal imbalance: prefrontal cortex (PFC) became hyperexcitable (glutamate release increased by 64.7%; sEPSC amplitude increased 2.05-fold, P<0.05), whereas the hippocampus showed synaptic suppression (GABA release reduced by 32.65%; sIPSC frequency reduced by 66.67%, P<0.05). Mechanistically, SCN2B overexpression suppressed BDNF/TrkB/PI3K/AKT signaling axis (BDNF reduced by 53.17%; p-TrkB reduced by 63.62%, P<0.05), accompanied by decreased synaptic proteins (PSD95 43.45%, synapsin I 40.31%, P<0.05). Remarkably, NICD-reduced Navβ2[V173A,V175K] mice were fully resistant to CUMS-induced behavioral and neurotransmitter deficits, establishing NICD as a critical mediator of stress vulnerability. Collectively, these findings unravel a causal pathway in which SCN2B overexpression drives NICD-dependent suppression of BDNF signaling, leading to region-specific excitatory/inhibitory imbalance, synaptic disintegration, and depression-like phenotypes. Targeting Navβ2 proteolysis to block NICD generation may thus represent a novel therapeutic strategy for mood disorders.

Epilepsy is a neurological condition that affects approximately 50 million individuals worldwide. Current treatments mainly alleviate symptoms and have substantial adverse effects, with about 1/3 of patients not responding to existing medications. This underscores the urgent need for new drugs that can overcome these challenges. According to recent studies, the Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway play an important role in epileptogenesis. Brain-derived neurotrophic factor (BDNF) and c-fos are also involved in neuronal excitability and seizure-induced neuronal changes. The purpose of the present study was to examine the anticonvulsant and antiepileptogenic activities of 3,4-dimethoxyacetophenone (3,4-DMA) in the acute seizure and pentylenetetrazole (PTZ)-induced kindling models of epilepsy and to elucidate related molecular changes of BDNF, c-fos, and JAK-2/STAT-3 signaling pathways. 3,4-DMA was first evaluated in an acute seizure model, followed by a kindling model of epilepsy in mice. The effects of 3,4-DMA on BDNF, c-fos, and JAK-2/STAT-3 were assessed by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and immunohistochemistry (IHC). The results showed that 3,4-DMA significantly reduced seizure scores in a PTZ-induced kindling model. BDNF, c-fos, and JAK-2/STAT-3 were significantly upregulated in both the cortical and hippocampal tissues in PTZ-control mice. Conversely, treatment with 3,4-DMA attenuated this overexpression in the cortex and hippocampus. In conclusion, 3,4-DMA has antiepileptogenic effect as evidenced by a reduction in the expression of BDNF, c-fos and the JAK-2/STAT-3 pathway components. So, it can be a promising therapeutic candidate in the treatment of epilepsy. However, future studies measuring p-JAK-2 and p-STAT-3 will be required to confirm pathway activation status.

Serotonin (5-HT) 2A receptors (5-HT2AR) modulate corticolimbic circuits underlying fear extinction, but the behavioral consequences of 5-HT2AR antagonism during extinction remain poorly defined. Here, we examined the systemic effects of the 5-HT2A/2C antagonist MDL 11,939 (Glemanserin) and the selective 5-HT2A antagonist MDL 100,907 (Volinanserin) on fear extinction in adult female and male C57BL/6J mice. Prior to drug administration, mice acquired comparable conditioned freezing responses during delay fear conditioning. Twenty-four hours later, acute administration of MDL 11,939 (1.0 mg/kg) or MDL 100,907 (0.01 mg/kg) increased freezing to the first conditioned stimulus (CS) presentation on Extinction Day 1 in males, indicating enhanced expression of conditioned fear. Repeated administration of MDL 11,939 significantly impaired extinction in male mice, evidenced by increased freezing across extinction trials and an increased number of trials required to reach extinction criterion. Neither acute nor repeated MDL 11,939 altered extinction in female mice. In males, MDL 100,907 produced a transient increase in freezing during extinction but did not delay extinction acquisition and had no effect in females. Because MDL 11,939 has reported affinity for 5-HT2C receptors, we tested potential contributions of 5-HT2C receptor antagonism in a separate cohort of male mice given repeated administration of the selective 5-HT2C antagonist SB 242084 (0.25 or 0.5 mg/kg). Neither dose affected conditioned fear expression, extinction learning, or trials required to reach extinction criterion. Together, these findings demonstrate ligand-, sex-, and dose-dependent effects of 5-HT2AR antagonism on fear extinction and suggest that distinct intracellular receptor signaling mechanisms may differentially regulate extinction-related behavior.

Breast cancer (BC) is the leading cause of cancer death among women worldwide with estrogen receptor positive (ER+) BC comprising 70-80% of all cases. Adjuvant therapy with aromatase inhibitors (AIs), such as letrozole (LTZ), is the standard of care in ER + BC patients to prevent recurrence but many patients cease treatment prematurely due to the development of musculoskeletal side effects (i.e. arthralgia, myalgia, decreased grip strength) referred to as aromatase inhibitor-associated musculoskeletal syndrome (AIMSS). The lack of efficacious treatments available for AIMSS and premature treatment cessation being linked to increased patient mortality highlights the critical importance of identifying new therapeutic targets for its management. Here, we directly link LTZ treatment with increased levels of sphingosine-1-phosphate receptor 1 (S1PR1) endogenous ligands in spinal cord tissue of female ovariectomized (OVX) ICR mice. We therefore investigated the impact of reducing S1PR1 activity as a potential therapeutic strategy for alleviating AIMSS-related behaviors. In C57BL/6J OVX female mice, we found that pharmacological inhibition using the selective and competitive S1PR1 antagonist, NIBR-0213, effectively alleviated AIMSS-like behaviors after acute and repeated dosing regimens, with limited therapeutic effectiveness when given prior to LTZ treatment. In a conditioned place preference (CPP) paradigm, LTZ- but not vehicle-treated animals preferred the NIBR-0213 paired chamber, unmasking the presence of a tonic-aversive state driven by LTZ treatment. These findings identify S1PR1 as an important mediator to AIMSS and suggest that S1PR1 antagonists may be most effective as acute interventions to provide relief from the sensory and affective burden of AIMSS.

Glucagon-like peptide-1 (GLP-1) receptor agonists have emerged as promising pharmacotherapeutics for alcohol use disorder (AUD). Here, we examined the effects of semaglutide, a long-acting GLP-1 receptor agonist, on alcohol intake and anxiety-related behaviors in Marchigian Sardinian alcohol-preferring rats, a model characterized by excessive alcohol intake and pronounced anxious and depressive-like phenotypes. Male and female rats were treated with semaglutide (0.001, 0.01 and 0.1 mg/kg, s.c.) and vehicle. Voluntary alcohol consumption was assessed using the two-bottle choice paradigm. Locomotion, anxiety, and stress responses were evaluated during withdrawal and in naïve rats, following administration of the highest dose of semaglutide (0.1 mg/kg). Furthermore, the acute effects of semaglutide on GABAergic transmission were assessed in the central nucleus of the amygdala and in the infralimbic (IL) medial pre-frontal cortex (mPFC) of naïve rats. Semaglutide reduced voluntary alcohol consumption in a dose-dependent manner, and it produced anxiolytic-like behaviors during withdrawal in the open field test. In naïve animals, semaglutide reduced locomotor activity, lowered blood glucose levels, and decreased fat mass and GABAergic transmission in the IL mPFC. Our findings support the use of the GLP-1 receptor agonist semaglutide as a promising pharmacotherapy for the treatment of AUD and its associated psychiatric symptoms.

Glioblastoma multiforme (GBM) is a WHO grade IV brain tumor with aggressive growth and frequent recurrence, and temozolomide (TMZ) remains the standard first-line chemotherapy despite limited durable benefits. Parthenolide (Pt), a sesquiterpene lactone derived from the medicinal plant feverfew, has shown antitumor activity in multiple cancer types; however, its therapeutic potential and downstream mechanisms in GBM have not been fully defined. Herein, we demonstrate that Pt induces apoptosis, suppresses stemness, and reduces invasion in human U-87MG and T98G cells. Pt also potentiates TMZ's efficacy in vitro and in an orthotopic GBM model with male NOD-SCID mice. RNA-sequencing and pathway analyses revealed that Pt markedly represses epithelial-mesenchymal transition (EMT)-associated programs. Among the most significantly downregulated genes, transgelin-2 (TAGLN2) emerged as a key Pt-responsive target, and its high expression was correlated with poor survival in glioma cohorts. TAGLN2 overexpression rescued Pt-mediated suppression of malignant phenotypes and the EMT, supporting a functional role for TAGLN2 in Pt responsiveness. Mechanistically, inhibitor screening identified glycogen synthase kinase-3β (GSK3β) as a critical upstream regulator of Pt-induced TAGLN2 repression. Promoter-reporter and ChIP-qPCR assays further showed that Pt activates GSK3β, leading to reduced cyclic AMP response-binding protein (CREB) transcriptional activity and diminished CREB occupancy at the TAGLN2 promoter, thereby suppressing TAGLN2 transcription. Collectively, our findings define a Pt-driven GSK3β/CREB/TAGLN2 axis that restrains the mesenchymal transition and enhances TMZ sensitivity in GBM, highlighting TAGLN2 and its upstream regulatory pathway as potential therapeutic targets for this lethal disease.

Binge drinking is a prevalent and clinically significant behavior contributing to alcohol use disorder. Sex differences in binge drinking are observed in both humans and preclinical studies, including differences in alcohol intake and underlying neural regulatory mechanisms. The cortical amygdala (CoA) has been implicated in alcohol-related behaviors, but its role in binge-like drinking and associated brain-wide co-activation patterns remains unclear. We tested whether CoA activity regulates binge-like alcohol consumption in a sex-dependent manner and examined CoA-associated brain-wide network organization in binge-like drinking. Binge-like alcohol intake was assessed using a 6-week Drinking-in-the-Dark paradigm. Saline or deschloroclozapine was administered 30 min before drinking in mice with inhibitory hM4Di, excitatory hM3Dq, or mCherry control virus. Locomotor and anxiety-like behavior were assessed 24 h after the final 4-h DID session. Brains were collected 120 min after drinking onset during a final DID session under chemogenetic manipulation. Whole-brain Fos mapping was performed, and brain-wide co-activation networks were constructed, followed by graph theory analyses. CoA inhibition significantly reduced binge-like alcohol consumption in male mice, but not females, although the drug × sex × virus interaction was not significant. CoA activation did not alter alcohol intake in either sex but increased locomotor activity in females but not males, although the corresponding sex interaction was not significant. Anxiety-like behavior was unaffected. CoA inhibition was associated with distinct brain-wide network configurations, including differences in CoA-associated module composition, hub-like node distribution, and co-activation patterns involving alcohol-relevant limbic and hypothalamic regions. These findings support a sex-dependent pattern of CoA regulation of binge-like drinking, with the reduction in alcohol intake detected in males but not females and demonstrate that CoA activity shapes brain-wide Fos co-activation patterns during alcohol consumption.

Prolonged social isolation triggers neuropsychiatric impairments linked to brain neurotransmitter imbalances. This study investigated the impact of social isolation on sensory perception, affective states, and the levels of amino acid neurotransmitters and endocannabinoids within the medial prefrontal cortex (mPFC) of both male and female adolescent mice. Furthermore, we evaluated the therapeutic efficacy of S-adenosylmethionine (SAMe), a methyl donor with established antidepressant properties, in mitigating these alterations. Single-housed (SH) male mice developed mechanical allodynia, cold hyperalgesia, depression-like behavior, and general malaise, alongside significantly decreased GABA levels in the mPFC. SAMe treatment effectively ameliorated these symptoms, restoring GABAergic tone and increasing glycine and glutamate levels. In contrast, SH female mice exhibited anxiety-like behavior and pain hypersensitivity without depression-like traits. Their neurochemical profile was characterized by decreased GABA but increased levels of glutamate and specific endocannabinoids/acylethanolamides (2-AG, PEA, and LEA) in the mPFC. In females, SAMe treatment successfully reversed anxiety-like behavior and pain hypersensitivity, normalized glutamate, LEA, and GABA levels, and increased glycine and DHG. Our findings demonstrate that while social isolation-induced neuropsychiatric and neurochemical alterations are sex-dependent, the development of sensory hypersensitivity and the therapeutic responsiveness to SAMe are consistent across sexes. By restoring GABAergic homeostasis and modulating excitatory and endocannabinoid signaling in the mPFC, SAMe emerges as a promising pharmacological intervention for treating the complex psychiatric and sensory sequelae induced by chronic social stress.

Postictal hypoxia causes several consequences that are detrimental to the quality of life of people with epilepsy. Cannabidiol (CBD) and Δ[9]-tetrahydrocannabinol (THC), the primary phytocannabinoids from cannabis, have been shown to exhibit vasoactive and antioxidant properties that may mitigate the local oxygen deficiency observed after seizures. Here, we focus on the effects of CBD and THC on seizure-induced hypoxia. Adult Long-Evans rats of both sexes were assigned to drug-specific cohorts and the effects on local hippocampal oxygen were measured before and after seizure elicitation following either vaporized or oral administration of the two phytocannabinoids. Acute effects were assessed following a single administration of each drug, whereas chronic effects were evaluated over 28 days of daily treatment. In naïve rats, acute vaporized CBD significantly attenuated postictal hypoxia in both sexes, whereas acute oral THC attenuated postictal hypoxia in male rats. With chronic daily administration, phytocannabinoids produced time-dependent adaptations. Chronic vaporized CBD in females and chronic oral THC in males maintained attenuating effects on postictal hypoxia but exhibited partial tolerance, characterized by diminished effects on postictal oxygen decline in both sexes, recovery of postictal oxygen in females, and severity of hypoxia in males. These results demonstrate that acutely and chronically administered phytocannabinoids attenuate postictal hypoxia in a route-, sex-, and time-dependent manner. Together, these findings provide evidence that vaporized CBD and oral THC hold potential to treat the postictal hypoxia-induced comorbidities associated with seizures, potentially increasing the overall quality of life of people with epilepsy.

Consolation, a prosocial behavior in which individuals comfort distressed conspecifics, is regarded as a fundamental expression of empathy. Orexin (OX) neurons in the lateral hypothalamus (LH) have been implicated in empathic processes, but the specific circuitry and receptor subtypes through which they modulate consolation remain unknown. Here, we investigated the role of LH[OX] neurons and their projection to the ventral tegmental area (VTA) in regulating consolation-like behavior in male mice. We found that consolation toward a defeat induced stressed partner was accompanied by reduced c-Fos expression in LH[OX] neurons. Using chemogenetic manipulations, we demonstrated that activation of LH[OX] neurons, or specifically the LH[OX]→VTA projections, decreased consolation-like allogrooming. Furthermore, intra-VTA injection of the orexin receptor 1 (OX1R) antagonist SB-334867, but not the OX2R antagonist Seltorexant, rescued the allogrooming deficit induced by pathway activation, identifying VTA OX1R as the critical receptor subtype. These effects occurred without alterations in anxiety-like behavior or general sociability. Collectively, these findings reveal that the LH[OX]→VTA pathway, acting through OX1R, contributes to the regulation of consolation, establishing a receptor-specific circuit mechanism for orexinergic modulation of empathy-related prosocial behavior.

Cerebral ischemic preconditioning (CIP) confers robust protection against subsequent ischemic stroke by enhancing brain ischemic tolerance. Although ferroptosis is increasingly recognized as a critical pathogenic mechanism in stroke, the role of N[6]-methyladenosine (m[6]A) RNA modification in its regulation remains poorly understood. This study investigated the involvement of m[6]A signaling in the anti-ferroptotic mechanism of CIP using complementary in vivo mouse middle cerebral artery occlusion (MCAO) and in vitro oxygen-glucose deprivation (OGD) models. We found that CIP significantly attenuated ischemia-induced neuronal ferroptosis in vivo, whereas OGD preconditioning (OGDP) recapitulated these anti-ferroptotic effects in HT22 cells. Acyl-CoA synthetase long-chain family member 4 (ACSL4), a key executor of ferroptosis, was markedly upregulated following ischemic injury, whereas CIP effectively suppressed this increase. Similarly, OGDP attenuated OGD-induced ACSL4 upregulation in HT22 cells. Importantly, ACSL4 overexpression significantly attenuated the protective effects of OGDP against OGD-induced ferroptosis. Mechanistic analyses in HT22 cells revealed that OGD downregulated the m[6]A demethylase fat mass and obesity-associated protein (FTO) and upregulated m[6]A readers insulin-like growth factor 2 mRNA-binding proteins (IGF2BPs), including IGF2BP1 and IGF2BP3, thereby stabilizing ACSL4 mRNA and promoting ferroptosis. OGDP alleviated these effects by restoring FTO expression, reducing IGF2BP1 and IGF2BP3 levels, and enhancing m[6]A-dependent ACSL4 mRNA decay. Together, these findings demonstrate that CIP induces ischemic tolerance and attenuates ACSL4-associated ferroptosis. Mechanistic studies in neuronal cells further identify the m[6]A-dependent FTO-IGF2BP1/3-ACSL4 axis as a potential regulatory mechanism contributing to OGDP-induced ferroptosis resistance.

Preoperative sleep deprivation (SD) is known to exacerbate postoperative pain, but the underlying mechanisms remain unclear. Using a male mouse model of 48-h continuous SD followed by incisional surgery, we found that SD alone induced transient mechanical hyperalgesia, and when combined with surgery (SI group) it significantly aggravated and prolonged postoperative pain recovery until day 10. On postoperative day 7, SI mice showed marked downregulation of spinal dopamine D2 receptor (D2R) expression and decreased spinal dopamine concentration. D2R was predominantly expressed on spinal glutamatergic neurons. Overexpression of D2R on these neurons via intrathecal AAV injection effectively alleviated the prolonged hyperalgesia, normalizing pain sensitivity by day 7. Mechanistically, D2R downregulation led to increased phosphorylation of NMDA receptor subunits (p-NR1 and p-NR2B) without altering total NMDA receptor levels. Co-immunoprecipitation revealed an interaction between D2R and protein phosphatase 2A (PP2A). SI mice exhibited decreased PP2A expression but increased p-AKT, p-GSK3β, and β-catenin, indicating activation of the PP2A/AKT signaling pathway, which were reversed by D2R overexpression. Furthermore, pharmacological activation of PP2A with DT-061 not only downregulated p-AKT, p-GSK3β, β-catenin, p-NR1 and p-NR2B, but also produced significant analgesia in SI mice. Collectively, these results demonstrate that preoperative sleep deprivation prolongs postoperative hyperalgesia by downregulating spinal D2R on glutamatergic neurons, which disinhibits the PP2A/AKT pathway and enhances NMDA receptor phosphorylation. Targeting D2R or its downstream PP2A/AKT signaling may represent a novel therapeutic strategy for managing persistent postoperative pain caused by preoperative sleep deprivation in male mice.