Synthesis and biological evaluation of radioiodinated benzoxazole and benzothiazole derivatives for imaging myelin in multiple sclerosis.

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system that results from destruction of the myelin sheath. Due to heterogeneity of the symptoms and course of MS, periodic monitoring of disease activity is important for diagnosis and treatment. In the present study, we synthesized four radioiodinated benzoxazole (BO) and benzothiazole (BT) derivatives, and evaluated their utility as novel myelin imaging probes for single photon emission computed tomography (SPECT). In a biodistribution study using normal mice, three compounds ([125I]BO-1, [125I]BO-2, and [125I]BT-2) displayed moderate brain uptake (2.7, 2.9, and 2.8% ID/g, respectively) at 2 min postinjection. On ex vivo autoradiography using normal mice, [125I]BO-2 showed the most preferable ratio of radioactivity accumulation in white matter (myelin-rich region) versus gray matter (myelin-deficient region). In addition, the radioactivity of [125I]BO-2 was reduced in the lysophosphatidylcholine-induced demyelination region. In conclusion, [123I]BO-2 demonstrated the fundamental characteristics of a myelin imaging probe for SPECT.

Wild-Type DCTN1 Suppresses the Aggregation of DCTN1 Mutants Associated with Perry Disease.

Perry disease, a rare autosomal dominant neurodegenerative disorder, is characterized by parkinsonism, depression or apathy, unexpected weight loss, and central hypoventilation. Genetic analyses have revealed a strong association between point mutations in the dynactin I gene (DCTN1) coding p150glued and Perry disease. Although previous reports have suggested a critical role of p150glued aggregation in Perry disease pathology, whether and how p150glued mutations affect protein aggregation is not fully understood. In this study, we comprehensively investigated the intracellular distribution of the p150glued mutants in HEK293T cells. We further assessed the effect of co-overexpression of the wild-type p150glued protein with mutants on the formation of mutant aggregates. Notably, overexpression of p150glued mutants identified in healthy controls, which is also associated with amyotrophic lateral sclerosis, showed a thread-like cytoplasmic distribution, similar to the wild-type p150glued. In contrast, p150glued mutants in Perry disease and motor neuron disease caused aggregation. In addition, the co-overexpression of the wild-type protein with p150glued mutants in Perry disease suppressed aggregate formation. In contrast, the p150glued aggregation of motor neuron disease mutants was less affected by the wild-type p150glued. Further investigation of the mechanism of aggregate formation, contents of the aggregates, and biological mechanisms of Perry disease could help develop novel therapeutics.

Amelioration of obsessive-compulsive disorder by intracellular acidification of cortical neurons with a proton pump inhibitor.

Obsessive-compulsive disorder (OCD) is a highly prevalent neuropsychiatric disorder poorly controlled with pharmacological treatment because of the wide variation in symptom patterns. We analysed real-world data on adverse self-reports and insurance claims to identify a novel therapeutic target for OCD. We found that dopamine D2 receptor (D2R) agonists increased the incidence of OCD-like symptoms, which were suppressed by the concomitant use of proton pump inhibitors (PPIs). Further, OCD-like repetitive and habitual behaviours were observed in mice repeatedly injected with a D2R agonist, quinpirole. However, these abnormalities were suppressed by short-term PPI treatment. In quinpirole-treated mice, PPI inhibited pyramidal neuron hyperactivity in the lateral orbitofrontal cortex, a region where the P-type proton pump gene Atp4a is abundantly expressed. In primary cultured cortical neurons, short-term PPI treatment lowered intracellular pH and decreased firing activity, which was mimicked by Atp4a knockdown. Our findings show that inhibition of P-type proton pumps may be a novel therapeutic strategy for OCD.

A Novel Strategy for the Discovery of Drug Targets: Integrating Clinical Evidence with Molecular Studies.

The mechanisms of several drugs remain unclear, limiting our understanding of how they exert their effects. Receptor affinities have not been comprehensively measured during drug development, and the safety investigations in humans are limited. Therefore, numerous unknown adverse and beneficial effects of drugs in humans persist. In this review, I highlight our achievements in identifying the unexpected beneficial effects of drugs through the analysis of real-world clinical data, which can contribute to drug repositioning and target finding. (1) Through the analysis of real-world data, we found that the anti-arrhythmic amiodarone induced interstitial lung disease, leading to fibrosis. Surprisingly, concurrent use of an anti-thrombin drug, dabigatran mitigated these adverse events. Pharmacological studies using animal models have mimicked this phenomenon and revealed the molecular mechanisms associated with the platelet-derived growth factor-alpha receptors. (2) The antidiabetic dipeptidyl-peptidase 4 inhibitors increased the risk of an autoimmune disease, bullous pemphigoid, which was reduced by the concomitant use of lisinopril. Pharmacological studies using human peripheral blood mononuclear cells have revealed that lisinopril suppressed the skin disorders by inhibiting the expression of cutaneous matrix metalloproteinase 9 in macrophages. (3) The antimicrobial fluoroquinolones increased the risk of tendinopathy, which was reduced by the concomitant use of dexamethasone. However, clinical guidelines have suggested that corticosteroid increases the risk of tendinopathy. Our investigation demonstrated that fluoroquinolones impaired tendon cells through DNA damage by generating reactive oxygen species. In contrast, dexamethasone exhibited an acute beneficial effect on tendon tissue by upregulating the expression of a radical scavenger, glutathione peroxidase 3.

Inhibitors of the Mechanistic Target of Rapamycin Can Ameliorate Bortezomib-Induced Peripheral Neuropathy.

Bortezomib, an anticancer drug for multiple myeloma and mantle cell lymphoma, causes severe adverse events and leads to peripheral neuropathy. The associated neuropathy limits the use of bortezomib and could lead to discontinuation of the treatment; therefore, effective intervention is crucial. In the present study, we statistically searched for a drug that could alleviate bortezomib-induced peripheral neuropathy using adverse event self-reports. We observed that specific inhibitors of the mechanistic target of rapamycin (mTOR) lowered the incidence of bortezomib-induced peripheral neuropathy. These findings were experimentally validated in mice, which exhibited long-lasting mechanical hypersensitivity after repeated bortezomib treatment. This effect was inhibited for hours after a systemic injection with rapamycin or everolimus in a dose-dependent manner. Bortezomib-induced allodynia was accompanied by the activation of spinal astrocytes, and intrathecal injection of mTOR inhibitors or an inhibitor of ribosomal protein S6 kinase 1, a downstream target of mTOR, exhibited considerable analgesic effects in a dose-dependent manner. These results suggest that mTOR inhibitors, which are readily available to patients prescribed bortezomib, are one of the most effective therapeutics for bortezomib-induced peripheral neuropathy.

The astrocytic TRPA1 channel mediates an intrinsic protective response to vascular cognitive impairment via LIF production.

Vascular cognitive impairment (VCI) refers to cognitive alterations caused by vascular disease, which is associated with various types of dementia. Because chronic cerebral hypoperfusion (CCH) induces VCI, we used bilateral common carotid artery stenosis (BCAS) mice as a CCH-induced VCI model. Transient receptor potential ankyrin 1 (TRPA1), the most redox-sensitive TRP channel, is functionally expressed in the brain. Here, we investigated the pathophysiological role of TRPA1 in CCH-induced VCI. During early-stage CCH, cognitive impairment and white matter injury were induced by BCAS in TRPA1-knockout but not wild-type mice. TRPA1 stimulation with cinnamaldehyde ameliorated BCAS-induced outcomes. RNA sequencing analysis revealed that BCAS increased leukemia inhibitory factor (LIF) in astrocytes. Moreover, hydrogen peroxide-treated TRPA1-stimulated primary astrocyte cultures expressed LIF, and culture medium derived from these cells promoted oligodendrocyte precursor cell myelination. Overall, TRPA1 in astrocytes prevents CCH-induced VCI through LIF production. Therefore, TRPA1 stimulation may be a promising therapeutic approach for VCI.

Infection-induced extracellular vesicles evoke neuronal transcriptional and epigenetic changes.

Infection with the protozoan Toxoplasma gondii induces changes in neurotransmission, neuroinflammation, and behavior, yet it remains elusive how these changes come about. In this study we investigated how norepinephrine levels are altered by infection. TINEV (Toxoplasma-induced neuronal extracellular vesicles) isolated from infected noradrenergic cells down-regulated dopamine ß-hydroxylase (DBH) gene expression in human and rodent cells. Here we report that intracerebral injection of TINEVs into the brain is sufficient to induce DBH down-regulation and distrupt catecholaminergic signalling. Further, TINEV treatment induced hypermethylation upstream of the DBH gene. An antisense lncRNA to DBH was found in purified TINEV preparations. Paracrine signalling to induce transcriptional gene silencing and DNA methylation may be a common mode to regulate neurologic function.

Increased expression of glutathione peroxidase 3 prevents tendinopathy by suppressing oxidative stress.

Tendinopathy, a degenerative disease, is characterized by pain, loss of tendon strength, or rupture. Previous studies have identified multiple risk factors for tendinopathy, including aging and fluoroquinolone use; however, its therapeutic target remains unclear. We analyzed self-reported adverse events and the US commercial claims data and found that the short-term use of dexamethasone prevented both fluoroquinolone-induced and age-related tendinopathy. Rat tendons treated systemically with fluoroquinolone exhibited mechanical fragility, histological change, and DNA damage; co-treatment with dexamethasone attenuated these effects and increased the expression of the antioxidant enzyme glutathione peroxidase 3 (GPX3), as revealed via RNA-sequencing. The primary role of GPX3 was validated in primary cultured rat tenocytes treated with fluoroquinolone or H2O2, which accelerates senescence, in combination with dexamethasone or viral overexpression of GPX3. These results suggest that dexamethasone prevents tendinopathy by suppressing oxidative stress through the upregulation of GPX3. This steroid-free approach for upregulation or activation of GPX3 can serve as a novel therapeutic strategy for tendinopathy.

Vitamin D supplementation is effective for olanzapine-induced dyslipidemia.

Olanzapine is an atypical antipsychotic drug that is clinically applied in patients with schizophrenia. It increases the risk of dyslipidemia, a disturbance of lipid metabolic homeostasis, usually characterized by increased low-density lipoprotein (LDL) cholesterol and triglycerides, and accompanied by decreased high-density lipoprotein (HDL) in the serum. In this study, analyzing the FDA Adverse Event Reporting System, JMDC insurance claims, and electronic medical records from Nihon University School of Medicine revealed that a co-treated drug, vitamin D, can reduce the incidence of olanzapine-induced dyslipidemia. In the following experimental validations of this hypothesis, short-term oral olanzapine administration in mice caused a simultaneous increase and decrease in the levels of LDL and HDL cholesterol, respectively, while the triglyceride level remained unaffected. Cholecalciferol supplementation attenuated these deteriorations in blood lipid profiles. RNA-seq analysis was conducted on three cell types that are closely related to maintaining cholesterol metabolic balance (hepatocytes, adipocytes, and C2C12) to verify the direct effects of olanzapine and the functional metabolites of cholecalciferol (calcifediol and calcitriol). Consequently, the expression of cholesterol-biosynthesis-related genes was reduced in calcifediol- and calcitriol-treated C2C12 cells, which was likely to be mediated by activating the vitamin D receptor that subsequently inhibited the cholesterol biosynthesis process via insulin-induced gene 2 regulation. This clinical big-data-based drug repurposing approach is effective in finding a novel treatment with high clinical predictability and a well-defined molecular mechanism.

Dorsal raphe serotonergic neurons preferentially reactivate dorsal dentate gyrus cell ensembles associated with positive experience.

Major depressive disorder (MDD) is among the most common mental illnesses. Serotonergic (5-HT) neurons are central to the pathophysiology and treatment of MDD. Repeatedly recalling positive episodes is effective for MDD. Stimulating 5-HT neurons of the dorsal raphe nucleus (DRN) or neuronal ensembles in the dorsal dentate gyrus (dDG) associated with positive memories reverses the stress-induced behavioral abnormalities. Despite this phenotypic similarity, their causal relationship is unclear. This study revealed that the DRN 5-HT neurons activate dDG neurons; surprisingly, this activation was specifically observed in positive memory ensembles rather than neutral or negative ensembles. Furthermore, we revealed that dopaminergic signaling induced by activation of DRN 5-HT neurons projecting to the ventral tegmental area mediates an increase in active coping behavior and positive dDG ensemble reactivation. Our study identifies a role of DRN 5-HT neurons as specific reactivators of positive memories and provides insights into how serotonin elicits antidepressive effects.

Early Detection of Adverse Drug Reaction Signals by Association Rule Mining Using Large-Scale Administrative Claims Data.

INTRODUCTION: Adverse drug reactions (ADRs) are a leading cause of mortality worldwide and should be detected promptly to reduce health risks to patients. A data-mining approach using large-scale medical records might be a useful method for the early detection of ADRs. Many studies have analyzed medical records to detect ADRs; however, most of them have focused on a narrow range of ADRs, limiting their usefulness. OBJECTIVE: This study aimed to identify methods for the early detection of a wide range of ADR signals. METHODS: First, to evaluate the performance in signal detection of ADRs by data-mining, we attempted to create a gold standard based on clinical evidence. Second, association rule mining (ARM) was applied to patient symptoms and medications registered in claims data, followed by evaluating ADR signal detection performance. RESULTS: We created a new gold standard consisting of 92 positive and 88 negative controls. In the assessment of ARM using claims data, the areas under the receiver-operating characteristic curve and the precision-recall curve were 0.80 and 0.83, respectively. If the detection criteria were defined as lift > 1, conviction > 1, and p-value < 0.05, ARM could identify 156 signals, of which 90 were true positive controls (sensitivity: 0.98, specificity: 0.25). Evaluation of the capability of ARM with short periods of data revealed that ARM could detect a greater number of positive controls than the conventional analysis method. CONCLUSIONS: ARM of claims data may be effective in the early detection of a wide range of ADR signals.

Peripheral Beta-2 Adrenergic Receptors Mediate the Sympathetic Efferent Activation from Central Nervous System to Splenocytes in a Mouse Model of Fibromyalgia.

Abnormalities in the peripheral immune system are involved in the pathophysiology of fibromyalgia, although their contribution to the painful symptoms remains unknown. Our previous study reported the ability of splenocytes to develop pain-like behavior and an association between the central nervous system (CNS) and splenocytes. Since the spleen is directly innervated by sympathetic nerves, this study aimed to examine whether adrenergic receptors are necessary for pain development or maintenance using an acid saline-induced generalized pain (AcGP) model (an experimental model of fibromyalgia) and whether the activation of these receptors is also essential for pain reproduction by the adoptive transfer of AcGP splenocytes. The administration of selective ß2-blockers, including one with only peripheral action, prevented the development but did not reverse the maintenance of pain-like behavior in acid saline-treated C57BL/6J mice. Neither a selective α1-blocker nor an anticholinergic drug affects the development of pain-like behavior. Furthermore, ß2-blockade in donor AcGP mice eliminated pain reproduction in recipient mice injected with AcGP splenocytes. These results suggest that peripheral ß2-adrenergic receptors play an important role in the efferent pathway from the CNS to splenocytes in pain development.

Specific Cellular Effects of Low Bortezomib Concentrations on Purified Cultures of Schwann Cells, Satellite Glial Cells, Macrophages, and Dorsal Root Ganglion Neurons.

Peripheral neuropathy is one of the major adverse effects that limit the clinical application of bortezomib (BTZ). However, the underlying mechanisms of BTZ-induced peripheral neuropathy (BIPN) remain elusive. To examine cell types potentially involved in the development of BIPN, we used four purified cultures of cells of the peripheral nervous system: Schwann cells (SCs), satellite glial cells (SGCs), macrophages, and dorsal root ganglion (DRG) neurons. Administration of a low BTZ concentration (5 nM; similar to concentrations in clinical use) caused dedifferentiation of cultured SCs, returning mature SCs to an immature state. In cultured SGCs, BTZ increased glial fibrillary acidic protein (GFAP) levels without inducing the release of inflammatory cytokines or chemokines. In macrophages, BTZ caused little inflammatory response. Finally, in DRG neurons, BTZ strongly suppressed the expression levels of sensor and transducer ion channels without affecting cell morphology. Taken together, low concentrations of BTZ can cause SC dedifferentiation (i.e., demyelination), increased GFAP level in SGC, and decreased expression levels of sensor and transducer ion channels in DRG neurons (i.e., numbness feeling). Thus, we have reported, for the first time, specific effects of BTZ on peripheral nervous system cells, thereby contributing to a better understanding of the initiating mechanism of BIPN.

Acetaminophen improves tardive akathisia induced by dopamine D2 receptor antagonists.

Tardive akathisia is a movement disorder characterized by internal restlessness with an uncontrollable urge to move, leading to repetitive movements. It is a common side effect of long-term treatment with dopamine D2 receptor antagonists. In the present study, we analyzed the FDA Adverse Event Reporting System and IBM MarketScan Research Database to find a drug that can be used concomitantly with dopamine D2 receptor antagonists and still reduce the risk of akathisia. Acetaminophen was determined to be the most effective akathisia-suppressing drug. In an experimental validation of the hypothesis, chronic treatment of rats with haloperidol caused akathisia symptoms, including increased stereotyped behavior and locomotor activity, and decreased immobility time. Acute treatment with acetaminophen significantly attenuated haloperidol-induced akathisia. In the ventral striata of these rats, acetaminophen prevented haloperidol-induced decrease in the number of c-Fos+ preproenkephalin+ neurons. These results suggest that acetaminophen is effective in suppressing tardive akathisia by activating indirect-pathway medium spiny neurons.

Median raphe serotonergic neurons projecting to the interpeduncular nucleus control preference and aversion.

Appropriate processing of reward and aversive information is essential for survival. Although a critical role of serotonergic neurons in the dorsal raphe nucleus (DRN) in reward processing has been shown, the lack of rewarding effects with selective serotonin reuptake inhibitors (SSRIs) implies the presence of a discrete serotonergic system playing an opposite role to the DRN in the processing of reward and aversive stimuli. Here, we demonstrated that serotonergic neurons in the median raphe nucleus (MRN) of mice process reward and aversive information in opposite directions to DRN serotonergic neurons. We further identified MRN serotonergic neurons, including those projecting to the interpeduncular nucleus (5-HTMRN→IPN), as a key mediator of reward and aversive stimuli. Moreover, 5-HT receptors, including 5-HT2A receptors in the interpeduncular nucleus, are involved in the aversive properties of MRN serotonergic neural activity. Our findings revealed an essential function of MRN serotonergic neurons, including 5-HTMRN→IPN, in the processing of reward and aversive stimuli.

Analysis of real-world data and a mouse model indicates that pirfenidone causes pellagra.

Background: Pirfenidone (PFD) is widely used in patients with idiopathic pulmonary fibrosis (IPF) and its adverse effects, such as nausea and photosensitivity, are well known. Many patients with IPF have reduced doses or even cessation of PFD because of its side-effects. No solutions have been found for these side-effects because the current mechanistic insights are insufficient. Methods: Using the results of real-world data analysis from the US Food and Drug Administration Adverse Events Reporting System, we hypothesised that PFD-related symptoms may be similar to pellagra. Reverse translational experiments using female BALB/c mice were performed to validate and estimate this hypothesis. Niacin and its metabolite responses were compared between patients with IPF treated with PFD and those treated without PFD. Results: The pellagra hypothesis was translated from real-world data analysis. Pharmacological and comprehensive genetic investigations showed that PFD caused pellagra-related nausea and photosensitivity in a mouse model, which may have been mediated by the actions of nicotinamide N-methyltransferase (NNMT). Higher NNMT substrate responses were observed in urine from patients and mice with PFD than in those without PFD. Conclusions: PFD may cause pellagra or pellagra-like symptoms such as photosensitivity. Further studies are required to investigate whether niacin prevents pellagra-like symptoms caused by PFD in patients with IPF.

Diff-ATAC-STARR-Seq: A Method for Genome-Wide Functional Screening of Enhancer Activity in Vivo.

Transcriptional regulatory elements, including promoters and enhancers, play a key role in the cell-type specific regulation of the transcriptome. Application of rapidly evolving genetic tools, such as optogenetic/chemogenetic actuators and fluorescent reporters to elucidate the function of cell subtypes in vivo necessitates cell-type specific promoters or enhancers. In this context, methods for genome-wide functional screening of cis-regulatory elements, including enhancers, are of utmost importance. In this study, we describe a novel method for genome-wide functional screening of enhancer activity in vivo with minimal handling. Application of the method to cells from different brain structures and subsequent differential analysis allow identification of active enhancers in the target tissue or brain structures. To demonstrate proof of concept, we applied this method to samples from the dorsal raphe nucleus (DRN) and the medial prefrontal cortex of the mouse brain and successfully identified six enhancers with highly biased activity towards the dorsal raphe nucleus. Considering that these two structures consist of largely similar cell types whereas serotonin and dopamine neurons exist only in the DRN, our results confirm the validity of this method in identifying cell-type specific and brain-structure specific enhancers. Overall, this method will be helpful in identifying cis-regulatory elements suitable for cell-type specific manipulations.

Autism Spectrum Disorder Model Mice Induced by Prenatal Exposure to Valproic Acid Exhibit Enhanced Empathy-Like Behavior via Oxytocinergic Signaling.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder characterized by core symptoms, including impairments in social behavior and repetitive interests. Recent studies have revealed that individuals with ASD also display decreased empathy, ultimately leading to difficulties in social relationships; however, another report indicated that individuals with ASD have enhanced emotional empathy. Nonetheless, the neurobiological mechanisms underlying altered empathy in individuals with ASD remain unclear. In this study, we assessed empathy-like behaviors in valproic acid (VPA)-treated mice-a mouse model of ASD with observational fear learning. We then investigated the brain regions and signaling systems responsible for the altered empathy-like behaviors in VPA-treated mice. As a result, mice prenatally exposed to VPA displayed increased empathy-like behaviors, which were not attributed to altered sensitivity to auditory stimuli or enhanced memory for pain-related contexts. Immunohistochemical analysis revealed that the number of c-Fos positive oxytocinergic neurons in the paraventricular nucleus of the hypothalamus (PVN) was significantly higher in VPA-treated mice after observational fear learning. Finally, we found that pretreatment with L-368899, an antagonist of the oxytocin receptor, repressed the empathetic behavior in VPA-treated mice. These results suggest that VPA-treated ASD model animals showed increased emotional empathy-like behaviors through the hyperactivation of PVN oxytocinergic neurons for the first time. Further investigation of this hyperactivity will help to identify extrinsic stimuli and the condition which are capable of activation of PVN oxytocinergic neurons and to identify novel approach to enhance oxytocin signaling, which ultimately pave the way to development of novel therapy for ASD.

Orai2 channel regulates prostaglandin E2 production in TNFα/IL1α-stimulated astrocytes.

Astrocytes are glial cells that serve homeostatic functions in the central nervous system (CNS). Recent research, however, suggests that under pathological conditions, astrocytes are stimulated by various factors and actively participate in CNS inflammation. In the present study, we found that astrocytes upregulate various inflammatory factors including prostaglandin E2 (PGE2 ) by co-stimulation with tumor necrosis factor-alpha (TNFα) and interleukin-1alpha (IL1α). These TNFα/IL1α-stimulated astrocytes also showed increased Ca2+ release from the endoplasmic reticulum (ER) and increased expression of Orai2, a member of the store-operated calcium channel (SOCC) family. To reveal the role of Orai2, we used astrocytes in which Orai2 was knocked-down (KD) or knocked-out (KO). The expression of the prostaglandin E synthase Ptges and the production of PGE2 were higher in Orai2-KD astrocytes than in WT astrocytes when stimulated with TNFα and IL1α. Orai2-KO astrocytes also showed increased expression of Ptges and increased PGE2 production. The expression of Ptgs2, another PGE2 synthetic enzyme, was also upregulated in Orai2-KO astrocytes. Moreover, Orai2-KO astrocytes showed increased store-operated calcium entry (SOCE) and increased Orai1 expression. These results suggest that Orai2 is upregulated in TNFα/IL1α-stimulated astrocytes and reduces PGE2 production to some extent, modulating CNS inflammation. Our findings may aid in understanding how astrocytes are associated with inflammatory responses, and the identification of new targets that modulate astrocytic reactivity.

Seasonal Prevalence of the Invasive Longhorn Beetle Aromia bungii in Osaka Prefecture, Japan.

A thorough understanding of the seasonal prevalence of invasive pests in newly invaded regions is key for establishing an appropriate and localized control plan for their successful eradication. In this study, we investigated the seasonal prevalence of the invasive longhorn beetle Aromia bungii (Coleoptera: Cerambycidae) in Osaka Prefecture, Japan. We determined the number of adult beetles sighted on host trees more than once a week from late May or early June to late August for 3 years at three study sites (one site from 2019-2021 and two sites from 2020-2021). The appearance period of A. bungii adults spanned over 2 months (June-August), and peak sighting in the field occurred in late June; the adults were more abundant in the early phase of their seasonal prevalence (around the peak dates) and almost disappeared by August. The number of adult beetles emerging from A. bungii-infested trees at one study site was surveyed daily in 2021. This survey showed a short-span adult emergence period: Approximately 1 month from the first emergence day, supporting the idea of the concentration of adult abundance in the early phase. These results will help to establish a timely pest-control plan for A. bungii in Osaka Prefecture.