Chronic migraine is a common central nervous system disorder characterized by recurrent, pulsating headaches. However, the extent and mechanisms of hypothalamic involvement in disease progression have not been thoroughly investigated. Herein, we created a chronic migraine mouse model using repeated intraperitoneal injections of nitroglycerin. We performed transcriptomic sequencing on the hypothalamus of mice with chronic migraine and control mice under normal physiological conditions, followed by differential gene set enrichment and functional analysis of the data. Additionally, we examined the intrinsic connection between chronic migraine and sleep disorders using transcriptomic sequencing data from sleep-deprived mice available in public databases. We identified 39 differentially expressed genes (DEGs) in the hypothalamus of a mouse model of chronic migraine. Functional analysis of DEGs revealed enrichment primarily in signaling transduction, immune-inflammatory responses, and the cellular microenvironment. A comparison of the transcriptomic data of sleep-deprived mice revealed two commonly expressed DEGs. Our findings indicate that the hypothalamic DEGs are primarily enriched in the PI3K/AKT/mTOR pathway and associated with the NF-kB/NLRP3/IL-1 β pathway activation to maintain the central sensitization of the chronic migraine. Chronic migraine-induced gene expression changes in the hypothalamus may help better understand the underlying mechanisms and identify therapeutic targets.

BACKGROUND: The periaqueductal gray (PAG) is at the center of a powerful descending antinociceptive neuronal network, and is a key node in the descending pain regulatory system of pain. However, less is known about the altered perfusion of PAG in chronic migraine (CM).
OBJECTIVE: To measure the perfusion of PAG matter, an important structure in pain modulation, in CM with magnetic resonance (MR) perfusion without contrast administration.
METHODS: Three-dimensional pseudocontinuous arterial spin labeling (3D-PCASL) and brain structure imaging were performed in 13 patients with CM and 15 normal subjects. The inverse deformation field generated by brain structure image segmentation was applied to the midbrain PAG template to generate individualized PAG. Then the perfusion value of the PAG area of the midbrain was extracted based on the individual PAG mask.
RESULTS: Cerebral blood flow (CBF) value of PAG in CM patients (47.98 ± 8.38 mL/100 mg min) was significantly lower than that of the control group (59.87 ± 14.24 mL/100 mg min). Receiver operating characteristic (ROC) curve analysis showed that the area under the curve was 0.77 (95% confidence interval [CI], 0.60, 0.94), and the cutoff value for the diagnosis of CM was 54.83 mL/100 mg min with a sensitivity 84.60% and a specificity 60%.
CONCLUSIONS: Imaging evidence of the impaired pain conduction pathway in CM may be related with the decreased perfusion in the PAG, which could be considered as an imaging biomarker for the diagnosis and therapy evaluation.

BACKGROUND: Migraine is a neurological disorder characterized by complex, widespread, and sudden attacks with an unclear pathogenesis, particularly in chronic migraine (CM). Specific brain regions, including the insula, amygdala, thalamus, and cingulate, medial prefrontal, and anterior cingulate cortex, are commonly activated by pain stimuli in patients with CM and animal models. This study employs fluorescence microscopy optical sectioning tomography (fMOST) technology and AAV-PHP.eB whole-brain expression to map activation patterns of brain regions in CM mice, thus enhancing the understanding of CM pathogenesis and suggesting potential treatment targets.
METHODS: By repeatedly administering nitroglycerin (NTG) to induce migraine-like pain in mice, a chronic migraine model (CMM) was established. Olcegepant (OLC) was then used as treatment and its effects on mechanical pain hypersensitivity and brain region activation were observed. All mice underwent mechanical withdrawal threshold, light-aversive, and elevated plus maze tests. Viral injections were administered to the mice one month prior to modelling, and brain samples were collected 2 h after the final NTG/vehicle control injection for whole-brain imaging using fMOST.
RESULTS: In the NTG-induced CMM, mechanical pain threshold decreased, photophobia, and anxiety-like behavior were observed, and OLC was found to improve these manifestations. fMOST whole-brain imaging results suggest that the isocortex-cerebral cortex plate region, including somatomotor areas (MO), somatosensory areas (SS), and main olfactory bulb (MOB), appears to be the most sensitive area of activation in CM (P < 0.05). Other brain regions such as the inferior colliculus (IC) and intermediate reticular nucleus (IRN) were also exhibited significant activation (P < 0.05). The improvement in migraine-like symptoms observed with OLC treatment may be related to its effects on these brain regions, particularly SS, MO, ansiform lobule (AN), IC, spinal nucleus of the trigeminal, caudal part (Sp5c), IRN, and parvicellular reticular nucleus (PARN) (P < 0.05).
CONCLUSIONS: fMOST whole-brain imaging reveals c-Fos + cells in numerous brain regions. OLC improves migraine-like symptoms by modulating brain activity in some brain regions. This study demonstrates the activation of the specific brain areas in NTG-induced CMM and suggests some regions as a potential treatment mechanism according to OLC.

Exosomes have been identified as optimal biomarkers to screen for multiple diseases. However, few studies focus on the abundant exosome population isolated from plasma of migraine. This study investigated whether proteins in abundant exosomes can aid in the diagnosis of chronic migraine (CM). Plasma exosomes were collected by centrifugation, from which protein samples were extracted. A pilot study (CM, 18; episodic migraine (EM), 26) followed by a second dataset (CM, 26; EM, 16; tension-type headache (TTH), 20; control, 22) was applied to establish a diagnostic model of CM. We employed proteomics based on liquid chromatography-tandem mass spectrometry (LC-MS/MS) to search for potential candidate biomarkers in plasma exosomes from CM patients. In total, 530 proteins in plasma exosomes were co-detected. Among them, 13 proteins were found significantly dysregulated between the plasma exosomes of CM patients and other groups. The receiver operating characteristic curve analysis revealed a combination of six proteins (upregulated: RAP2B, AK1, BID, DAG1, PICALM, PSMB2) could distinguish CM patients with high accuracy. Linear correlation analysis showed that the combination was significantly correlated with Headache Impact Test (HIT-6) scores (assessing the negative impact of headaches on normal daily activity). The RT-qPCR results showed the same trends in CM models with nitroglycerin as the exosomal protein sequencing results. These data revealed dysregulated proteins in plasma exosomes of CM, and the combination of plasma exosomal proteins RAP2B, AK1, BID, DAG1, PICALM, and PSMB2 could serve as a novel candidate biomarker for CM diagnosis.

To explore the mechanism of the classic formula Sanpian Decoction in treating chronic migraine, this study employed the four-dimensional data-dependent acquisition(4D-DIA) proteomics to analyze the effect of the decoction on chronic migraine in rats and experimentally verified the key differentially expressed proteins. Firstly, SD male rats were randomly divided into groups and repeatedly injected with nitroglycerin to prepare a chronic migraine model. After 7 consecutive days of gavage, rat grimace scale(RGS) was employed to evaluate the treatment efficacy. The trigeminal ganglion was collected for 4D-DIA proteomics, on the basis of which the diffe-rentially expressed proteins between groups were screened. Multiple databases were used for the Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment of the differentially expressed proteins. STRING and Cytoscape were employed to establish the protein-protein interaction(PPI) network. Western blot was employed to determine the expression level of the key diffe-rentially expressed protein TRPV1. The results showed that there were 517 differentially expressed proteins between blank group and model group and 221 differentially expressed proteins between model group and medium-dose Sanpian Decoction group. The GO and KEGG enrichment results showed that these differentially expressed proteins were mainly related to inflammatory response, injurious sensory stimulation, triglyceride metabolism, immune regulation, etc., which mainly involved the inflammation-related TRP, AMPK, PI3K-Akt, and TGF-β signaling pathways. The PPI network showed that the target proteins such as IGF, TOP2A, APOA1, CDK1, TTN, RYR1, and CSRP3 had high degrees. Compared with that in model group, the expression level of TRPV1 altered in medium-and high-dose Sanpian Decoction group(P<0.05). In conclusion, Sanpian Decoction may treat chronic migraine by regulating the inflammation-related pathways such as TRP, AMPK, and PI3K-Akt. It plays an important role in the regulation of TRPV1 protein and potentially modulates the perception of injurious stimuli, lipid metabolism, and immune responses.

BACKGROUND: Alpha 7 nicotinic acetylcholine receptor (α7nAChR)-mediated astrocytic activation is closely related to central sensitization of chronic migraine (CM). Xiongzhi Dilong decoction (XZDL), originated from Xiongzhi Shigao decoction of Yi-zong-jin-jian, has been confirmed to relieve CM in experiment and clinic. However, its underlying mechanism for treating CM has not been elucidated.
OBJECTIVE: To reveal the underlying mechanisms of XZDL to alleviate CM in vivo focusing mainly on α7nAChR-mediated astrocytic activation and central sensitization in TNC.
METHODS: CM rat model was established by subcutaneous injection of nitroglycerin (NTG) recurrently, and treated with XZDL simultaneously. Migraine-like behaviors of rats (ear redness, head scratching, and cage climbing) and pain-related reactions (mechanical hind-paw withdrawal threshold) of rats were evaluated before and after NTG injection and XZDL administration at different points in time for nine days. The immunofluorescence single and double staining were applied to detect the levels of CGRP, c-Fos, GFAP and α7nAChR in NTG-induced CM rats. ELISA kits were employed to quantify levels of TNF-α, IL-1β, and IL-6 in medulla oblongata of CM rats. The expression levels of target proteins were examined using western blotting. Finally, methyllycaconitine citrate (MLA, a specific antagonist of α7nAChR) was applied to further validate the mechanisms of XZDL in vivo.
RESULTS: XZDL significantly attenuated the pain-related behaviors of the NTG-induced CM rats, manifesting as constraints of aberrant migraine-like behaviors including elongated latency of ear redness and decreased numbers of head scratching and cage climbing, and increment of mechanical withdrawal threshold. Moreover, XZDL markedly lowered levels of CGRP and c-Fos, as well as inflammatory cytokines (IL-1β, IL-6 and TNF-α) in CM rats. Furthermore, XZDL significantly enhanced α7nAChR expression and its co-localization with GFAP, while markedly inhibited the expression of GFAP and the activation of JAK2/STAT3/NF-κB pathway in the TNC of CM rats. Finally, blocking α7nAChR with MLA reversed the effects of XZDL on astrocytic activation, central sensitization, and the pain-related behaviors in vivo.
CONCLUSIONS: XZDL inhibited astrocytic activation and central sensitization in NTG-induced CM rats by facilitating α7nAChR expression and suppressing JAK2/STAT3/NF-κB pathway, implying that the regulation of α7nAChR-mediated astrocytic activation represents a novel mechanism of XZDL for relieving CM.

OBJECTIVE: Chronic migraine is a disabling progressive disorder without effective management approaches. Animal models have been developed and used in chronic migraine research. However, there are several problems with existing models. Therefore, we aimed to summarize and analyze existing animal models to facilitate translation from basic to clinical.
RESULTS: The most commonly used models are the inflammatory soup induction model and the nitric oxide donor induction model. In addition, KATP openers have also been used in model induction. Based on the above models, some molecular targets have been identified, such as glutamate receptors. However, each model has its shortcomings and characteristics, and there are still some common problems that need to be solved, such as spontaneous headache, evaluation criteria after model establishment, and identification methods. In this review, we summarized and highlighted the advantages and limitations of the currently commonly used animal models of chronic migraine with a special focus on drug discovery and current therapeutic strategies, and discussed the directions that can be worked on in the future.

BACKGROUND: Chronic migraine (CM) significantly impacts both the physical and mental health of patients. Current studies on the safety and effectiveness of different pharmacological prophylaxis interventions for CM are limited. To address this gap, we conducted a network meta-analysis (NMA) to compare and rank the efficacy and safety of various drugs in preventing CM.
METHODS: Two independent researchers systematically searched four databases from their inception to August 1, 2023, to identify eligible randomized controlled trials (RCTs). Subsequently, they performed data extraction and assessed the risk of bias. A NMA was then performed. Continuous outcomes and binary outcomes were displayed as weighted mean difference (WMD) and risk ratio (RR), respectively, and corresponding 95% confidence intervals (CI) were reported. The surface under the cumulative ranking curve (SUCRA) was used to rank each intervention separately.
RESULTS: 24 RCTs involving 8789 patients were included. Compared to placebo, Botulinum toxin A demonstrated the most significant effect in reducing the monthly migraine days for CM patients (MD = 3.88, 95% CI 0.48, 7.28); in terms of improving the response rate by a 50% reduction in monthly migraine days, Topiramate (RR = 50.06, 95% CI 3.18, 787.30) was the most effective; there was no statistically significant difference between all preventive drugs and placebo in improving the migraine disability assessment (MIDAS) score; in terms of the incidence of adverse events, Eptinezumab (RR = 1.09, 95% CI 0.8, 1.54) exhibited the highest safety profile.
CONCLUSIONS: Among all the drugs for the preventive drugs for CM, Botulinum toxin A has the best efficacy and safety profile, closely followed by calcitonin gene-related peptide (CGRP) monoclonal antibodies (mAbs).

UNASSIGNED: Central sensitization is one of the pivotal pathological mechanisms in chronic migraine (CM). Silent information regulator 1 (SIRT1) was shown to be involved in CM, but its specific mechanism is unclear. Reactive oxygen species (ROS) are increasingly regarded as important signaling molecules in several models of pain. However, studies about the role of ROS in the central sensitization of CM model are rare. We thus explored the specific process of SIRT1 involvement in the central sensitization of CM, focusing on the ROS pathway.
UNASSIGNED: Inflammatory soup was repeatedly administered to male Sprague-Dawley rats to establish a CM model. The SIRT1 expression level in trigeminal nucleus caudalis (TNC) tissues was assessed by qRT-PCR and Western blotting analysis. The levels of ROS were detected by a Tissue Reactive Oxygen Detection Kit, DHE staining, and the fluorescence signal intensity of 8-OHdG. A ROS scavenger (tempol), a SIRT1 activator (SRT1720), a SIRT1 inhibitor (EX527), and a mitochondrial fission inhibitor (Mdivi-1) were used to investigate the specific molecular mechanisms involved. NMDAR2B, CGRP, ERK, and mitochondrial fission-related protein were evaluated by Western blotting, and the CGRP level in frozen sections of the TNC was detected via immunofluorescence staining.
UNASSIGNED: After repeated inflammatory soup infusion and successful establishment of the CM rat model, SIRT1 expression was found to be significantly reduced, accompanied by elevated ROS levels. Treatment with Tempol, SRT1720, or Mdivi-1 alleviated allodynia and reduced the increase in NMDAR2B phosphorylation and CGRP and ERK phosphorylation in the CM rat. In contrast, EX527 had the opposite effect in CM rat. SRT1720 and EX527 decreased and increased ROS levels, respectively, in CM rats, and tempol reversed the aggravating effect of EX527 in CM rats. Furthermore, the regulatory effect of SIRT1 on ROS may include the involvement of the mitochondrial fission protein DRP1.
UNASSIGNED: The results indicate the importance of SIRT1 in CM may be due to its role in regulating the production of ROS, which are involved in modulating central sensitization in CM. These findings could lead to new ideas for CM treatment with the use of SIRT1 agonists and antioxidants.

OBJECTIVE: To compare the real-world effectiveness and tolerability of calcitonin gene-related peptide (CGRP) monoclonal antibodies (mAbs) and onabotulinumtoxinA in chronic migraine (CM) patients.
METHODS: This multicenter study involved retrospective analysis of prospectively collected data of CM patients treated with CGRP mAbs or onabotulinumtoxinA, including difficult-to-treat (DTT) patients (i.e., ≥3 preventive failures). Treatment outcomes were determined at 6 months based on prospective headache diaries and Migraine Disability Assessment (MIDAS).
RESULTS: The study included 316 (55 M/261F, mean age 44.4 ± 13.5 years) and 333 (61 M/272F, mean age 47.9 ± 13.4 years) CM patients treated with CGRP mAbs or onabotulinbumtoxinA, respectively. At 6 months, CGRP mAb treatment was associated with a greater decrease in monthly migraine days (MMDs) (-13.0 vs. -8.7 days/month, p < 0.001) and a higher ≥50% responder rate (RR) (74.7% vs. 50.7%, p < 0.001) compared with onabotulinumtoxinA injections. The findings were consistent in DTT patients (-13.0 vs. -9.1 MMDs, p < 0.001; ≥50% RR: 73.9% vs. 50.3%, p < 0.001) or those with medication-overuse headache (MOH) (-13.3 vs. -9.0 MMDs, p < 0.001; ≥50% RR: 79.0% vs. 51.6%, p < 0.001). Besides, patients receiving CGRP mAbs had greater improvement (-42.2 vs. -11.8, p < 0.001) and a higher ≥50% RR (62.0% vs. 40.0%, p = 0.001) in MIDAS scores and a lower rate of adverse events (AEs) (6.0% vs. 21.0%, p < 0.001). However, none of the patients discontinued treatment due to AEs.
CONCLUSIONS: In this multicenter, real-world study, CGRP mAbs were more effective than onabotulinumtoxinA in CM patients, even in DTT or MOH patients. All of these injectables were well tolerated. Further prospective studies are needed to verify these findings.