{"paper_id":"2cbefd47-3ea8-4c66-889b-6fe648e0f59a","body_text":"Effects of Sumatriptan on PACAP, PAC-1, CGRP, VIP, and TRPV-1 Molecules in an Experimental Migraine Model | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Effects of Sumatriptan on PACAP, PAC-1, CGRP, VIP, and TRPV-1 Molecules in an Experimental Migraine Model Fatma ÖZTÜRK KÜP, Seher YILMAZ, Aslı OKAN, Sümeyye UÇAR, Ece EROĞLU, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7785868/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 13 You are reading this latest preprint version Abstract Migraine is a chronic neurological disorder characterized by severe headache. In the pathology of migraine, it is suggested that increased parasympathetic output to intracranial arteries due to excessive stimulation of the trigeminovascular system is responsible. The release of PACAP, VIP, and CGRP signaling molecules due to excessive stimulation leads to the dilation of intracranial arteries, causing headaches. Sumatriptan exerts its therapeutic effect by causing vasoconstriction of dilated meningeal blood vessels in migraine and suppressing the release of vasoactive neuropeptides from trigeminal sensory neurons. However, the relationship between the Sumatriptan agent and the signaling pathways in migraine pathology has not been sufficiently studied. In this study, we aimed to determine the potential therapeutic efficacy of the Sumatriptan agent on PACAP, PAC-1, CGRP, VIP, and TRPV1 molecules in an experimental migraine model. In the study, 62 Sprague Dawley rats (31 males and 31 females) weighing 250–300 grams were randomly divided into five experimental groups: Control, Sham, Migraine, Sumatriptan, and Migraine-Sumatriptan, with procedures followed accordingly. At the end of the study, histopathological evaluation of brain tissues taken from the animals was performed, and the immunoreactivity of PACAP, PAC-1, CGRP, VIP, and TRPV1 proteins was assesse immunohistochemically. As a result, it was observed that the histopathological findings of the sumatriptan treatment group were reduced compared to the migraine groups. Based on the obtained data, it was determined that sumatriptan significantly affected the neuropeptides associated with pain in migraine. However, more detailed analyses are still needed in this regard. Migraine Neurovascular Theory Nitroglycerin Sumatriptan Trigeminal neurons Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 INTRODUCTION Migraine, the second leading cause of disability worldwide, is a common neurological disorder affecting approximately 12% of the population (Stovner et al., 2007 ; Burch et al., 2019 ). Migraine is defined as a recurrent primary headache disorder resulting in attacks lasting 4–72 hours. The headache seen in migraine is typically unilateral, episodic, of moderate or severe intensity, and can be aggravated by routine physical activity. Migraine attacks can be associated with nausea, photophobia, and phonophobia (Headache Classification Committee of the International Headache Society, 2013). Despite these data, while the role of the trigeminovascular system in the pathology of migraine is suggested, this condition is not yet fully understood (Ashina et al., 2019 ; Charles, 2018). In this context, it has been stated that the neurovascular theory induces functional and anatomical changes in the pathophysiology of migraine (Marichal-Cancino et al., 2021 ). In the development of migraine, excessive stimulation of the trigeminovascular system (Hoffman et al., 2019) results in parasympathetic output to the intracranial arteries. This output, in turn, leads to the dilation of intracranial arteries due to the release of signaling molecules such as pituitary adenylate cyclase-activating peptide (PACAP) and calcitonin gene-related peptide (CGRP) (Ashina et al., 2017 ). In this context, targeting signaling molecules like PACAP and CGRP in new approaches could have therapeutic importance. PACAP and its binding receptor, pituitary adenylate cyclase-activating polypeptide type 1 (PAC-1), belong to the vasoactive intestinal peptide (VIP)/secretin/growth hormone-releasing neuropeptide superfamily, which is widely expressed in tissues (Wong et al., 1998 ; Arimura, 2007 ). The existing literature suggests that PACAP and its receptors may play a role in the pathogenesis of migraine-associated headaches (Edvinsson et al., 2018 ). CGRP, a neuropeptide related to signaling pathways in migraine, is produced in peripheral and central neurons and exhibits a potent vasodilatory effect (Brain et al., 1985 ; Rosenfeld et al., 1983 ). In this context, drug trials targeting CGRP in migraine-related attacks have come to the forefront (Tepper, 2019 ). According to the presented data, signaling molecules such as PACAP, PAC-1, VIP, and CGRP stand out as critical molecules in the occurrence of episodic attacks in migraine disease. Additionally, molecules that increase pain sensitivity in migraine-related attacks may also contribute to the enhancement of symptoms. Transient receptor potential (TRP) channels, which are among the molecules that increase pain sensitivity in the perception of migraine-related attacks, consist of six subfamilies: TRPA, TRPC, TRPM, TRPP, TRPL, and TRPV. These channels are primarily expressed on the cell membrane and are part of the cation channel family (Jardín et al., 2017 ). TRP channels contribute to various physiological processes, including temperature regulation and the perception of pain. In this context, transient receptor potential vanilloid 1 (TRPV1) channels, particularly among the TRP channels, are associated with migraine pathogenesis due to the stimulation of meningeal nociceptors related to pain (Benemei and Dussor, 2019 ). Furthermore, the activation of TRP channels can positively affect the release of CGRP from sensory nerve endings (Dussor et al., 2014 ). Therefore, it can be considered that TRPV1, from the TRP channel family, may play a role in the pathophysiological mechanisms related to migraine. In light of all the presented data, targeting PACAP, CGRP, VIP, which cause vasodilation, and TRPV1 channels, which increase pain sensitivity, with potential pharmacological trials in migraine-related attacks may have a positive effect on the acute treatment of migraine. Sumatriptan, one of the drugs used in the acute treatment of migraine, is a 5-HT1 agonist that selectively targets the 5-HT1B and 5-HT1D receptors. Sumatriptan exerts its therapeutic effect in migraine by causing vasoconstriction of dilated meningeal blood vessels, suppressing the release of vasoactive neuropeptides from perivascular trigeminal sensory neurons, and reducing the transmission of pain signals in the trigeminal dorsal horn (Ferrari et al., 2002 ; Goadsby, 2007 ). In another study, the relationship of Sumatriptan with TRPV channels in trigeminal neurons was examined. It was shown that TRPV1 channels are functional in neurons projecting to the cerebral dura and that Sumatriptan suppresses TRPV1 in trigeminal neurons (Evans et al., 2012 ). TRPV1 channels may have a specific role in the pathology of migraine. Additionally, a study conducted in 2021 on 26 migraine patients indicated that early treatment with intravenously administered Sumatriptan prevented PACAP-38-induced migraine (Wienholtz et al., 2021 ). In light of these data, it can be stated that Sumatriptan may affect PACAP-38 in relation to the trigeminovascular system. In the study, the potential effects of Sumatriptan, an agent used in the acute treatment of migraine, on PACAP and its receptor PAC-1, CGRP and VIP, which modulates CGRP, and the TRPV1 channel, which increases pain sensitivity, were investigated, as these have been shown to be related to migraine pathogenesis in the literature. Following this, a treatment group given the Sumatriptan agent was formed, and the animals were dissected. In this context, the immunoreactivity of PACAP, PAC-1, CGRP, VIP, and TRPV1 was evaluated using the immunohistochemistry method. MATERIALS AND METHODS An experimental migraine animal model was created in the project and approval was received for the study from the Erciyes University Animal Experiments Local Ethics Committee (Ethics committee protocol number: 22/076). In the study, 62 Sprague dawley animals, 31 females and 31 males, weighing 250–300 g, were randomly divided into 5 experimental groups and the procedures were observed. During the experiment, rats were housed in cages with a constant ambient temperature (24 ± 2°C), humidity (60 ± 5%), and alternating 12-h light and 12-h dark cycles. Experimental groups were designed as follows. Control group (n = 10, 5 females, 5 males) Rats in this group were fed rodent rearing pellet with free access to food and water for 14 days. Sham group (n = 10, 5 females, 5 males) Rats were given 1 ml of saline solution (0.9%) intraperitoneally (i.p.) for 14 days. Nitroglycerin Migraine group (n = 14, 7 females, 7 males) Nitroglycerin agent was administered at 10 mg/kg i.p. to 14 Spraque dawley rats, 7 females and 7 males, in the nitroglycerin-induced migraine group for 14 days. Nitroglycerin agent was dissolved in 30% alcohol, 30% propylene glycol and water, and this solution was diluted in 0.9% saline in a polypropylene tube and injected i.p. into rats (Lai et al., 2019 ). Sumatriptan group (n = 14, 7 females, 7 males) Rats in this group were given sumatriptan i.p. at a dose of 600 µg/kg for 14 days (Kurul et al., 2008 ; Casili et al., 2020 ). Nitroglycerin Migraine + Sumatriptan group (n = 14, 7 females, 7 males) Rats in this group were given 10 mg/kg i.p. nitroglycerin agent for 14 days. Nitroglycerin agent was dissolved in 30% alcohol, 30% propylene glycol and water. This solution was diluted in 0.9% saline in a polypropylene tube and injected i.p. into rats, and 600 µg/kg sumatriptan agent was given i.p. 5 minutes after nitroglycerin induction (Lai et al., 2019 ; Kural et al., 2008; Casili et al., 2020 ). While the rats were under anesthesia, they were sacrificed with the exsanguination method and the hippocampus and cortex tissue were dissected (Doğanyigit et al., 2023; Hamamcı et al., 2020; Ozkul et al., 2020 ; Akyuz et al., 2021 ). Histological Analysis The tissues were fixed with 10% formaldehyde solution and then washed in tap water (1 night), then dehydration was performed by passing the tissues through increasing alcohol series (70%, 80%, 90% and 100%). Then, the tissue samples were left in xylene (1082984000, Merck) to make them transparent, and after passing through paraffin series, they were embedded in clean paraffin. 5 µm thick sections taken from paraffin-embedded brain tissue samples using a microtome were stained with Harris hematoxylin and eosin. Histomorphological examination was performed under a light microscope (Olympus BX53) and imaged with a ZEISS Axiscope Colibri 3 digital camera. Neuron degeneration and vascular dilatation in the hippocampus and cortex were evaluated in the brain tissues of the experimental groups. Eosinophilic neuron with pyknotic nuclei, cell swelling or shrinkage was considered as neuron degeneration (Tambe et al., 2016 ; Doğanyiğit et al., 2023 ). Histopathological results in each category were scored as 0 = none, 1 = mild, 2 = moderate and 3 = severe. Quantification was performed randomly and blindly by two researchers. A minimum of 10 images were used for histomorphological scoring for at least 10 rats (5 females + 5 males) per experimental group. Immunohistochemistry Analysis Anti-PACAP (Affinity, DF7197), Anti-PAC-1 (Bioss, bs-0198R), Anti-CGRP (Affinity, DF7277), Anti-VIP (Affinity, DF6627) and Anti-TRPV-1 (Cloud-Clone Corp., PAF839Hu01) immunoreactivities were detected by immunohistochemical analysis (Okan et al., 2024 ; Doğanyiğit et al., 2023 ). In summary, after deparaffinization of 5µm thick sections, citrate buffer was used to open the epitopes (pH: 6.0; Thermo Fischer Scientific, UK, AP-9003-500). The slides were then placed in a 3% hydrogen peroxide solution in methanol to prevent endogenous peroxidase activity. Ultra V block solution (Thermo Fischer Scientific, UK, TA-125-UB) was applied to prevent nonspecific staining. Then, it was incubated with the primary antibody at 4oC overnight. Then, it was incubated with biotinylated goat anti polyvalent secondary antibody (Thermo Fischer Scientific, UK, TP-125-BN) in an oven at 37 o C for 40 minutes. After washing several times with PBS, it was incubated with streptavidin peroxidase (Thermo Fischer Scientific, UK, TS-125-HR) in an oven at 37oC for 30 minutes. The antibody complex was visualized by incubation with diaminobenzidine (DAB) chromogen (Thermo Fischer Scientific, UK, TA-125-HD). Then, the sections were counterstained with Gill III Hematoxylin (Merck, Germany, 1.05174.1000). It was dehydrated by passing through an increasing series of alcohol and sealed with a sealer called Entellan. Sections were examined with a ZEISS Axiscope Colibri 3 light microscope. Assessment of immunoreactivity levels was performed with Image J Version 1.46 (National Institutes of Health, Bethesda, Maryland). Statistical analysis Data were expressed as mean ± SD and analyzed by One-way ANOVA test and Tukey's post-hoc test for parametric tests. In non-parametric tests, independent sample-t test with median minimum-maximum values will be applied. p < 0.05 was considered significant in the analyses. RESULTS Histological Findings Histopathological changes in the brain hippocampus and prefrontal cortex areas of the rats in the experimental groups were evaluated under a light microscope after hematoxylin and eosin staining (Fig. 1 ). When neuron degeneration in the hippocampus in the nitroglycerin groups was compared, it was observed that it was more severe in males than in females (***p = 0.0004). While vascular dilation increased in males compared to females in the sumatriptan groups (p = 0.0045), more dilated blood vessels were observed in females than males in the nitroglycerin groups (p < 0.0001). When the cortex was examined, it was found that there was no difference between the experimental groups in terms of neuron degeneration. Vascular dilatation in the cortex was observed to be greater in males than in females in the sumatriptan groups (p = 0.0223). Neuron degeneration and vascular dilatation in the brain were observed to be significantly increased in the nitroglycerin groups in both male and female groups (Table 1). It was determined that neuron degeneration in the female and male cortices was significantly reduced in the nitroglycerin + sumatriptan groups compared to the nitroglycerin groups (Table 1). It was observed that the vascular dilation observed in the hippocampus of nitroglycerin + sumatriptan females was less severe than that of nitroglycerin females (Table 1). Table 1. Histopathological damage rates in brain tissues of experimental groups. Hippocampus Cortex Groups Neuron Degeneration Vascular dilation Neuron Degeneration Vascular dilation Control female 0.3 ± 0.07 0.32 ± 0.13 0.34 ± 0.08 0.52 ± 0.19 Sham female 0.22 ± 0.08 0.28 ± 0.04 0.34 ± 0.15 0.5 ± 0.36 Sumatriptan female 0.41 ± 0.13 0.3 ± 0.14 0.45 ± 0.19 0.34 ± 0.16 Nitroglycerin female 1.37 ± 0.61 abc 1.77 ± 0.17 abc 2.12 ± 0.14 abc 1.55 ± 0.32 abc Nitroglycerin +Sumatriptan female 1.55 ± 0.19 abc 0.78 ± 0.19 abcd 1.52 ± 0.17 abcd 0.91 ± 0.31 cd Control male 0.18 ± 0.08 0.38 ± 0.30 0.4 ± 0.14 0.48 ± 0.29 Sham male 0.26 ± 0.05 0.28 ± 0.08 0.38 ± 0.13 0.42 ± 0.13 Sumatriptan male 0.54 ± 0.27 0.67 ± 0.38 f 0.3 ± 0.14 0.75 ± 0.22 Nitroglycerin male 1.98 ± 0.19 efg 0.88 ± 0.10 ef 2.18 ± 0.19 efg 1.39 ± 0.21 efg Nitroglycerin +Sumatriptan male 1.77 ± 0.24 efg 0.61 ± 0.08 f 1.7 ± 0.23 efgh 1.13 ± 0.30 ef Immunohistochemistry Findings TRPV1 expression in the hippocampus was lower in nitroglycerin and nitroglycerin + sumatriptan groups compared to control, sham and sumatriptan groups. In both nitroglycerin and nitroglycerin + sumatriptan groups, TRPV1 expression in the hippocampus was higher in females than in males (Fig. 2 , p < 0.0001). TRPV1 expression in the cortex was significantly different between males and females in sham (p < 0.0001), sumatriptan (p = 0.0403) and nitroglycerin + sumatriptan (p = 0.0459) groups (Fig. 2 ). In the cortex, TRPV1 expression was decreased in sumatriptan, nitroglycerin and nitroglycerin + sumatriptan groups in females and males compared to the control group (Fig. 2 , p < 0.05). When the control and sham groups were compared within themselves in terms of VIP expression, a difference was observed between female and male hippocampi. In female hippocampuses, VIP expression decreased in all groups except the control group, whereas in male hippocampuses, VIP expression increased in the nitroglycerin + sumatriptan group compared to the other groups (Fig. 3 ). When VIP expression in the cortex was compared between the control, sham and nitroglycerin groups, a difference was observed between male and female rats (Fig. 3 ). In the cortex, VIP expression was decreased in the other experimental groups compared to the control group in both females and males (Fig. 3 ). When the control groups were compared within themselves in the hippocampus, a difference was observed between male and female rats in terms of PACAP expression. In Nitroglycerin + Sumatriptan groups, increased PACAP expression was observed in both male and female rats in the hippocampus (Fig. 4 ). When the control, sham and sumatriptan groups were compared within themselves in the cortex, it was found that there was a difference between male and female rats in terms of PACAP expression. In both male and female rats, PACAP expression in the hippocampus increased in nitroglycerin + sumatriptan groups, whereas in the cortex it increased in nitroglycerin groups (Fig. 4 ). In the hippocampus, when the nitroglycerin group was compared for P1 expression in females and males, it was observed that P1 expression was higher in females than in males. Increased P1 expression was observed in the hippocampus and cortex of female nitroglycerin and nitroglycerin + sumatriptan groups. In addition, P1 expression was increased in male hippocampus in nitroglycerin + sumatriptan group and decreased in nitroglycerin only groups. In the cortex, P1 expression was observed to be different in control, sham, nitroglycerin and nitroglycerin + sumatriptan groups in males and females. In male nitroglycerin + sumatriptan groups, P1 expression in the cortex was significantly increased (Fig. 5 ). of the experimental groups. Black arrow indicates immunoreactive cells. Magnification 20X and scale bar 50 µm. B) Bar graphs showing the intensity of immunostaining. Two-way analysis of variance, Tukey's and Sidak's multiple comparison tests were applied. Data shown in the bar graph are expressed as mean ± SD. In the female experimental groups, a p<0.05 indicates a statistically significant difference compared to the control group, b p<0.05 compared to the sham group, c p<0.05 compared to the sumatriptan group, d p<0.05 compared to the nitroglycerin group. In the male experimental groups, e p<0.05 compared to the control group, f p<0.05 compared to the sham group, g p<0.05 compared to the sumatriptan group and h p<0.05 compared to the nitroglycerin group. In the hippocampus, when sumatriptan and nitroglycerin groups were compared within themselves, a difference was observed in terms of CGRP expression in males and females.CGRP expression was significantly increased in the sumatriptan and nitroglycerin sumatriptan groups compared to the control group in males and females (Fig. 6 ). When the control, sumatriptan and nitroglycerin + sumatriptan groups were compared within themselves in the cortex, a difference was observed in terms of CGRP expression in males and females. Compared to the control group, CGRP expression increased in female sumatriptan groups, while it decreased in males. CGRP expression was similar in female and male nitroglycerin groups. While CGRP expression decreased in female nitroglycerin + sumatriptan group compared to nitroglycerin group, CGRP expression increased in male nitroglycerin + sumatriptan groups compared to nitroglycerin group (Fig. 6 ). DISCUSSION In this study, the effect of sumatriptan on neuropeptides in migraine disease was examined immunoexpressively. Sumatriptan-induced changes in PACAP, PAC-1, CGRP, VIP and TRPV-1 molecules were evaluated immunohistochemically. In both male and female groups, neuronal degeneration and vascular dilatation in the brain were significantly increased in nitroglycerin groups. In addition, sumatriptan significantly decreased neuronal degeneration in the brain in the nitroglycerin groups. In line with the data obtained, it was determined that sumatriptan significantly affected the neuropeptides associated with pain in migraine. Although triptan agents effective in the treatment of acute migraine are well tolerated, the mechanism of action is still debated. In this context, the present study provides an approach that sumatriptan will affect neuropeptides in the brain in migraine disease. In the literature, the relationship between sumatriptan and TRPV1 channels in trigeminal neurons has been examined. It has been shown that TRPV1 channels are functional in neurons projecting to the cerebral dura and sumatriptan suppresses TRPV1 in trigeminal neurons (Evans et al., 2012 ). Our results revealed that TRPV1 immunoactivity in the hippocampus was lower in both the experimental migraine group and the sumatriptan-treated migraine group compared to the control group. These data were significantly higher in the female group compared to males. In the cortex, TRPV1 was significantly decreased in both sexes in the migraine group receiving sumatriptan, the migraine group and the migraine group receiving the sumatriptan agent compared to the control. In line with these data, TRPV1 molecule may play a role in the brain in migraine-induced craniofacial pain. Sumatriptan may be involved in the reduction of migraine-related pain by affecting TRPV channels at the molecular level. Progestogens, estrogens and androgens may play a role in affecting migraine-induced TRPV1 in sumatriptan treatment. TRPV expression may be transcriptionally regulated by sex hormones (Jung et al., 2009 ). Based on these data, the role of sex hormones in the activation, modulation and regulation of the main TRPV channels involved in migraine pathophysiology should be considered. Parasympathetic neurotransmitters, PACAP38 and VIP, can be released from parasympathetic fibers and activate sensory nerve fibers during migraine attacks. It is known that 5-HT1B/D receptor activation has no effect on circulating VIP and PACAP levels in humans without trigeminovascular activation (Hansen et al., 2013 ). In our study, VIP expression decreased in female hippocampi except for the control group, whereas VIP expression increased in male hippocampi in the nitroglycerin + sumatriptan group compared to the other groups. In the cortex, VIP expression decreased in the other experimental groups compared to the control group in both females and males. In the literature, it has been shown that 2-hour VIP infusion triggers migraine attacks in migraine patients without aura. VIP was found to have a migraine triggering rate of 71% (Pellesi et al., 2020 ). In this context, VIP values that differ from the literature should be re-evaluated in the attack pathology of migraine disease in the brain. Given that neuropeptides play an important role in migraine, it may be important to conduct a similar study in the migraine population. Experimental studies have shown that PACAP isoforms (PACAP27 and PACAP38) cause migraine attacks after 20 minutes of infusion in migraine patients without aura (Ghanizada et al., 2020 ). Our results revealed that PACAP immune reactivity was increased in the hippocampus in the sumatriptan-treated migraine group in both females and males. In the cortex, increased PACAP expression was observed in migraine groups of both sexes. Similar to the literature, the relationship between PACAP increase and migraine disease has been shown histopathologically. A study reported that early treatment with intravenously administered sumatriptan prevented migraine caused by PACAP-38 (Wienholtz et al., 2021 ). In 2022, it was reported that sumatriptan had no effect on PACAP38-induced hypersensitivity associated with migraine (Guo et al., 2022 ). In our results, increased PACAP expression was observed in the hippocampus of the migraine experimental group treated with sumatriptan. Therefore, the activity of sumatriptan on PACAP in the brain should be repeated in different groups. Our finding may be interpreted as additional evidence that PACAP-mediated hypersensitivity is different from the known pathways in migraine. Preclinical evidence suggests that PACAP38 modulates trigeminal nociceptive activity mainly through PAC1 receptors, while clinical studies report that plasma concentrations of PACAP38 are elevated in cluster headache and migraine spontaneous attacks and return to normal after sumatriptan treatment (Zagami et al., 2014 ). In our study, more PAC1 expression was observed in the hippocampus in females. There is insufficient information on gender-related PAC1 expression in the literature. In this context, PAC1 expression may be modulated in migraine due to gender differences and estrogen dominance. In addition, PAC1 expression increased in the hippocampus and cortex in female nitroglycerin and nitroglycerin + sumatriptan groups. P1 expression was found to be significantly increased in the cortex in male nitroglycerin + sumatriptan groups. In addition, PAC1 expression was increased in the male hippocampus in the nitroglycerin + sumatriptan group and decreased in the nitroglycerin only group. These data suggest that the efficacy of sumatriptan may vary in males and females. Our finding draws attention to gender differences in the mechanism of PAC1-mediated PACAP in the known pathways of sumatriptan activity in migraine. Sumatriptan does not alter cephalic and extrencephalic circulating CGRP in healthy volunteers (Hansen et al., 2009 ). In our study, in the experimental migraine model, CGRP expression was found to be significantly increased in the hippocampus in male and female sumatriptan and nitroglycerin sumatriptan groups compared to the control group. In the cortex, CGRP expression was increased in female sumatriptan groups and decreased in males compared to the control group. In the female nitroglycerin + sumatriptan group, CGRP expression decreased compared to the nitroglycerin group, while in the male nitroglycerin + sumatriptan groups, CGRP expression increased compared to the nitroglycerin group. In this context, the efficacy of sumatriptan on CGRP in migraine may vary depending on the gender parameter. Fluctuations in estrogen levels modulate CGRP receptor signaling in the trigeminovascular system (Labastida-Ramírez et al., 2019 ). The presence of sex hormone receptors in the trigeminovascular system may indicate that trigeminal neurons are sensitive to changes in the levels of these hormones (Gupta et al., 2011 ). Our study revealed that sumatriptan, which is used in migraine attacks and belongs to the triptan family, may modulate neuropeptides in the hippocampus and cortex that have been implicated in migraine pathology. Significant sex-related changes in CGRP and TRPV1 immuno expression may be due to endocrine factors such as estrogen hormone. Considering that estrogens modulate the action of nociceptive receptors, female mice may have a lower pain threshold than male mice (Mitrovic et al., 2003 ). However, it may also be important to evaluate the condition during menstruation in females in migraine pathology. The effect of sumatriptan on the menstrual period and migraine attack in females can be discussed in more detail. In pathologies with trigeminal activation such as migraine, 5-HT1B/D receptor activation such as sumatriptan may regulate neuronal VIP and PACAP. The effect of sumatriptan on PACAP- related sensitization in migraine should be evaluated. In our study, increased PACAP expression was observed in the hippocampus of the migraine experimental group treated with sumatriptan. In this context, the significant increase in neuronal PACAP expression may be considered as an effective factor on hypersensitivity. In addition, the mechanism of the PAC1- mediated PACAP-mediated PACAP activity in sumatriptan may also draw attention to gender differences in known pathways in migraine. These data suggest that 5HT1 agonists such as sumatriptan may mediate neuronal neuropeptide changes by regulating the activation of the trigeminal system. This potential therapeutic mechanism of sumatriptan can be evaluated in systems such as migraine or cluster headache to establish its relevance. On the other hand, more detailed analyses are needed to analyze all proteins associated with the trigeminal system, to show which molecules are involved in regulatory mechanisms and to suggest therapeutic approaches for these molecules. Similarly, it is important to reveal the data in this study with different detection methods such as western blot. Declarations Conflict of interest The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. Funding This project was funded by Erciyes University Scientific Research Projects Unit (ERU BAPSIS) (Project support code: FBA-2023-12615). Author Contribution Fatma ÖZTÜRK KÜP; Validation, Writing – original draft. Seher YILMAZ; Formal analysis; Data curation. Aslı OKAN; Formal analysis; Data curation. Sümeyye UÇAR; Methodology, Conceptualization. Ece EROĞLU; Methodology, Conceptualization. Şükrü ATEŞ; Methodology, Conceptualization. A.Cihangir UĞUZ; Methodology, Conceptualization. Züleyha DOĞANYİĞİT; Writing, review & editing. Acknowledgement We thank Erciyes University Scientific Research Projects Unit (ERU BAPSIS) for funding support for this project. References Akyuz, E., Doganyigit, Z., Paudel, Y. N., Koklu, B., Kaymak, E., Villa, C., Arulsamy, A., Shaikh, M. F., Devinsky, O. (2021). “Immunoreactivity of Muscarinic Acetylcholine M2 and Serotonin 5-HT2B Receptors, Norepinephrine Transporter and Kir Channels in a Model of Epilepsy”. Life (Basel, Switzerland), 11(4), 276. Arimura A. (2007). “PACAP: the road to discovery”. Peptides, 28(9), 1617–1619. Ashina, M., Hansen, J. M., Á Dunga, B. O., Olesen, J. (2017). “Human models of migraine - short-term pain for long-term gain”. Nature reviews. Neurology, 13(12), 713–724. Ashina, M., Hansen, J. M., Do, T. P., Melo-Carrillo, A., Burstein, R., Moskowitz, M. A. (2019). “Migraine and the trigeminovascular system-40 years and counting”. The Lancet. Neurology, 18(8), 795–804. Benemei, S., Dussor, G. (2019). “TRP channels and migraine: recent developments and new therapeutic opportunities”. Pharmaceuticals, 12(2), 54. Boni, L. J., Ploug, K. B., Olesen, J., Jansen-Olesen, I., Gupta, S. (2009). “The in vivo effect of VIP, PACAP-38 and PACAP-27 and mRNA expression of their receptors in rat middle meningeal artery”. Cephalalgia: an international journal of headache, 29(8), 837–847. Brain, S. D., Williams, T. J., Tippins, J. R., Morris, H. R., MacIntyre, I. (1985). “Calcitonin gene-related peptide is a potent vasodilator”. Nature, 313(5997), 54–56. Burch, R. C., Buse, D. C., Lipton, R. B. (2019). “Migraine: Epidemiology, Burden, and Comorbidity”. Neurologic clinics, 37(4), 631–649. Casili, G., Lanza, M., Filippone, A., Campolo, M., Paterniti, I., Cuzzocrea, S., Esposito, E. (2020). “Dimethyl fumarate alleviates the nitroglycerin (NTG)-induced migraine in mice”. Journal of neuroinflammation, 17(1), 1-Charles A. (2018). “The pathophysiology of migraine: implications for clinical management”. The Lancet. Neurology, 17(2), 174–182. Doğanyiğit Z, Okan A, Akyüz E, Yılmaz S, Ateş Ş, Taheri S, Yılmaz Z, Shaikh MF. (2023). “Can endoplasmic reticulum stress observed in the PTZ-kindling model seizures be prevented with TUDCA and 4-PBA?” Eur J Pharmacol. 2023;960:176072. 10.1016/j.ejphar.2023.176072 . Doğanyiğit, Z., Okan, A., Kaymak, E., Pandır, D., Silici, S. (2020). “Investigation of protective effects of apilarnil against lipopolysaccharide induced liver injury in rats via TLR 4/HMGB- 1/NF-κB pathway”. Biomedicine & Pharmacotherapy, 125, 109967. Dussor, G., Yan, J., Xie, J. Y., Ossipov, M. H., Dodick, D. W., Porreca, F. (2014). “Targeting TRP channels for novel migraine therapeutics”. ACS chemical neuroscience, 5(11), 1085–1096. Edvinsson, L., Tajti, J., Szalárdy, L., Vécsei, L. (2018). “PACAP and its role in primary headaches”. The journal of headache and pain, 19(1), 1–7. Erdling, A., Sheykhzade, M., Maddahi, A., Bari, F., Edvinsson, L. (2013). “VIP/PACAP receptors in cerebral arteries of rat: characterization, localization and relation to intracellular calcium”. Neuropeptides, 47(2), 85–92. Evans, M. S., Cheng, X., Jeffry, J. A., Disney, K. E., Premkumar, L. S. (2012). “Sumatriptan inhibits TRPV1 channels in trigeminal neurons”. Headache, 52(5), 773–784. Ferrari, M. D., Goadsby, P. J., Roon, K. I., Lipton, R. B. (2002). “Triptans (serotonin, 5- HT1B/1D agonists) in migraine: detailed results and methods of a meta-analysis of 53 trials”. Cephalalgia, 22(8), 633–658. Gallai, V., Alberti, A., Gallai, B., Coppola, F., Floridi, A., Sarchielli, P. (2003). “Glutamate and nitric oxide pathway in chronic daily headache: evidence from cerebrospinal fluid”. Cephalalgia: an international journal of headache, 23(3), 166–174. Ghanizada, H., Al-Karagholi, M. A., Arngrim, N., Olesen, J., Ashina, M. (2020). “PACAP27 induces migraine-like attacks in migraine patients”. Cephalalgia: an international journal of headache, 40(1), 57–67. Goadsby, P. J. (2007). “Recent advances in understanding migraine mechanisms, molecules and therapeutics”. Trends in molecular medicine, 13(1), 39–44. Goadsby, P. J., Edvinsson, L., Ekman, R. (1988). “Release of vasoactive peptides in the extracerebral circulation of humans and the cat during activation of the trigeminovascular system”. Annals of neurology, 23(2), 193–196. Guo, S., Ernstsen, C., Hay-Schmidt, A., Kristensen, D. M., Ashina, M., Olesen, J., Christensen, S. L. (2022). “PACAP signaling is not involved in GTN-and levcromakalim-induced hypersensitivity in mouse models of migraine”. The Journal of Headache and Pain, 23(1), 155. Guo, Z., Czerpaniak, K., Zhang, J., Cao, Y. Q. (2021). “Increase in trigeminal ganglion neurons that respond to both calcitonin gene-related peptide and pituitary adenylate cyclase-activating polypeptide in mouse models of chronic migraine and posttraumatic headache”. Pain, 162(5), 1483–1499. Gupta, S., McCarson, K. E., Welch, K. M. A., Berman, N. E. (2011). “Mechanisms of pain modulation by sex hormones in migraine”. Headache: The Journal of Head and Face Pain, 51(6), 905–922. Hamamci, M., Doganyigit, Z., Silici, S., Okan, A., Kaymak, E., Yilmaz, S., Tokpinar, A., Inan, L. E. (2020). “Apilarnil: A Novel Neuroprotective Candidate”. Acta neurologica Taiwanica, 29(2), 33–45. Hanci, F., Kilinc, Y. B., Kilinc, E., Turay, S., Dilek, M., Kabakus, N. (2021). “Plasma levels of vasoactive neuropeptides in pediatric patients with migraine during attack and attack-free periods”. Cephalalgia: an international journal of headache, 41(2), 166–175. Hansen, J. M., Fahrenkrug, J., Petersen, J., Wienecke, T., Olsen, K. S., Ashina, M. (2013). “Vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) in the circulation after sumatriptan”. Scandinavian journal of pain, 4(4), 211–216. Hansen, J. M., Petersen, J., Wienecke, T., Olsen, K. S., Jensen, L. T., Ashina, M. (2009). “Sumatriptan does not change calcitonin gene-related peptide in the cephalic and extracephalic circulation in healthy volunteers”. The Journal of headache and pain, 10, 85–91.). Headache Classification Committee of the International Headache Society (IHS) (2013). “The International Classification of Headache Disorders, 3rd edition (beta version)”. Cephalalgia: an international journal of headache, 33(9), 629–808. Hoffmann, J., Baca, S. M., Akerman, S. (2019). “Neurovascular mechanisms of migraine and cluster headache”. Journal of cerebral blood flow and metabolism: official journal of the International Society of Cerebral Blood Flow and Metabolism, 39(4), 573–594. Jardín, I., López, J. J., Diez, R., Sánchez-Collado, J., Cantonero, C., Albarrán, L., Woodard, G. E., Redondo, P. C., Salido, G. M., Smani, T., Rosado, J. A. (2017). “TRPs in Pain Sensation”. Frontiers in physiology, 8, 392. Jung, C., Fandos, C., Lorenzo, I. M., Plata, C., Fernandes, J., Gené, G. G., Valverde, M. A. (2009). “The progesterone receptor regulates the expression of TRPV4 channel”. Pflügers Archiv-European Journal of Physiology, 459, 105–113. Kurul, S. H., Demirpence, S., Kiray, M., Tugyan, K., Yilmaz, O., Kose, G. (2008). “Investigation of the immunoreactivities of NOS enzymes and the effect of sumatriptan in adolescent rats using an experimental model of migraine”. The journal of headache and pain, 9(5), 317–323. Labastida-Ramírez, A., Rubio-Beltrán, E., Villalón, C. M., MaassenVanDenBrink, A. (2019). “Gender aspects of CGRP in migraine”. Cephalalgia, 39(3), 435–444. Lai, T., Chen, L., Chen, X., He, J., Lv, P., Ge, H. (2019). “Rhynchophylline attenuates migraine in trigeminal nucleus caudalis in nitroglycerin-induced rat model by inhibiting MAPK/NF-кB signaling”. Molecular and cellular biochemistry, 461(1–2), 205–212. Liu, J., Wang, G., Dan, Y., Liu, X. (2022). “CGRP and PACAP-38 play an important role in diagnosing pediatric migraine”. The journal of headache and pain, 23(1), 68. Marichal-Cancino, B. A., González-Hernández, A., Guerrero-Alba, R., Medina-Santillán, R., Villalón, C. M. (2021). “A critical review of the neurovascular nature of migraine and the main mechanisms of action of prophylactic antimigraine medications”. Expert review of neurotherapeutics, 21(9), 1035–1050. Mitrovic, I., Margeta-Mitrovic, M., Bader, S., Stoffel, M., Jan, L. Y., Basbaum, A. I. (2003). “Contribution of GIRK2-mediated postsynaptic signaling to opiate and alpha 2-adrenergic analgesia and analgesic sex differences”. Proceedings of the National Academy of Sciences of the United States of America, 100(1), 271–276. Okan, A., Demir, N., & Doğanyiğit, Z. (2024). “Linagliptin in combination with insülin suppresses apoptotic unfolded protein response in ovaries exposed to type 1 diabetes”. Cell biochemistry and function, 42(2), e3898. Ozkul, Y., Taheri, S., Bayram, K. K., Sener, E. F., Mehmetbeyoglu, E., Öztop, D. B., Aybuga, F., Tufan, E., Bayram, A., Dolu, N., Zararsiz, G., Kianmehr, L., Beyaz, F., Doganyigit, Z., Cuzin, F., Rassoulzadegan, M. (2020). “A heritable profile of six miRNAs in autistic patients and mouse models”. Scientific reports, 10(1), 9011. Pellesi, L., Al-Karagholi, M. A., Chaudhry, B. A., Lopez, C. L., Snellman, J., Hannibal, J., Amin, F. M., Ashina, M. (2020). “Two-hour infusion of vasoactive intestinal polypeptide induces delayed headache and extracranial vasodilation in healthy volunteers”. Cephalalgia: an international journal of headache, 40(11), 1212–1223. Pérez-Pereda, S., Toriello-Suárez, M., Ocejo-Vinyals, G., Guiral-Foz, S., Castillo-Obeso, J., Montes-Gómez, S., Martínez-Nieto, R. M., Iglesias, F., González-Quintanilla, V., Oterino, A. (2020). “Serum CGRP, VIP, and PACAP usefulness in migraine: a case-control study in chronic migraine patients in real clinical practice”. Molecular biology reports, 47(9), 7125–7138. Rosenfeld, M. G., Mermod, J. J., Amara, S. G., Swanson, L. W., Sawchenko, P. E., Rivier, J., Vale, W. W., Evans, R. M. (1983). “Production of a novel neuropeptide encoded by the calcitonin gene via tissue-specific RNA processing”. Nature, 304(5922), 129–135. Shibata, M., Tang, C. (2021). “Implications of transient receptor potential cation channels in migraine pathophysiology”. Neuroscience bulletin, 37(1), 103–116. Stovner, L. j., Hagen, K., Jensen, R., Katsarava, Z., Lipton, R., Scher, A., Steiner, T., Zwart, J. A. (2007). “The global burden of headache: a documentation of headache prevalence and disability worldwide”. Cephalalgia: an international journal of headache, 27(3), 193–210. Syed, A. U., Koide, M., Braas, K. M., May, V., Wellman, G. C. (2012). “Pituitary adenylate cyclase-activating polypeptide (PACAP) potently dilates middle meningeal arteries: implications for migraine”. Journal of molecular neuroscience: MN, 48(3), 574–583. Takács-Lovász, K., Kun, J., Aczél, T., Urbán, P., Gyenesei, A., Bölcskei, K., Szőke, É., Helyes, Z. (2022). “PACAP-38 Induces Transcriptomic Changes in Rat Trigeminal Ganglion Cells Related to Neuroinflammation and Altered Mitochondrial Function Presumably via PAC1/VPAC2 Receptor-Independent Mechanism”. International journal of molecular sciences, 23(4), 2120. Tambe, R., Jain, P., Patil, S., Ghumatkar, P., Sathaye, S. (2016). “Antiepileptogenic effects of borneol in pentylenetetrazole-induced kindling in mice”. Naunyn-Schmiedeberg's archives of pharmacology, 389(5), 467–475. Tepper S. J. (2019). “CGRP and headache: a brief review”. Neurological sciences: official journal of the Italian Neurological Society and of the Italian Society of Clinical Neurophysiology, 40 (Suppl 1), 99–105. Togha, M., Ghorbani, Z., Ramazi, S., Zavvari, F., Karimzadeh, F. (2021). “Evaluation of Serum Levels of Transient Receptor Potential Cation Channel Subfamily V Member 1, Vasoactive Intestinal Polypeptide, and Pituitary Adenylate Cyclase-Activating Polypeptide in Chronic and Episodic Migraine: The Possible Role in Migraine Transformation”. Frontiers in neurology, 12, 770980. Wienholtz, N., Christensen, C. E., Zhang, D. G., Coskun, H., Ghanizada, H., Al-Karagholi, M. A., Hannibal, J., Egeberg, A., Thyssen, J. P., Ashina, M. (2021). “Early treatment with an international journal of headache, 41(6), 731–748. Wong, A. O., Leung, M. Y., Shea, W. L., Tse, L. Y., Chang, J. P., Chow, B. K. (1998). “Hypophysiotropic action of pituitary adenylate cyclase-activating polypeptide (PACAP) in the goldfish: immunohistochemical demonstration of PACAP in the pituitary, PACAP stimulation of growth hormone release from pituitary cells, and molecular cloning of pituitary type I PACAP receptor”. Endocrinology, 139(8), 3465–3479. Zagami, A. S., Edvinsson, L., Goadsby, P. J. (2014). “Pituitary adenylate cyclase activating polypeptide and migraine”. Annals of clinical and translational neurology, 1(12), 1036–1040. Additional Declarations No competing interests reported. Supplementary Files GraphicalAbstract.jpg EthicalStatement.docx Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Revision requested 27 Nov, 2025 Reviews received at journal 25 Nov, 2025 Reviews received at journal 11 Nov, 2025 Reviewers agreed at journal 29 Oct, 2025 Reviewers agreed at journal 22 Oct, 2025 Reviewers agreed at journal 22 Oct, 2025 Reviewers agreed at journal 22 Oct, 2025 Reviewers agreed at journal 20 Oct, 2025 Reviewers agreed at journal 07 Oct, 2025 Reviewers invited by journal 07 Oct, 2025 Editor assigned by journal 06 Oct, 2025 Submission checks completed at journal 06 Oct, 2025 First submitted to journal 05 Oct, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {\"props\":{\"pageProps\":{\"initialData\":{\"identity\":\"rs-7785868\",\"acceptedTermsAndConditions\":true,\"allowDirectSubmit\":false,\"archivedVersions\":[],\"articleType\":\"Research Article\",\"associatedPublications\":[],\"authors\":[{\"id\":531190102,\"identity\":\"c99d2221-17dd-4f1e-8e42-20560cb38935\",\"order_by\":0,\"name\":\"Fatma ÖZTÜRK KÜP\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Erciyes University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Fatma\",\"middleName\":\"ÖZTÜRK\",\"lastName\":\"KÜP\",\"suffix\":\"\"},{\"id\":531190104,\"identity\":\"dd1f07c6-90e1-4979-974a-a1da1cad3534\",\"order_by\":1,\"name\":\"Seher YILMAZ\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Yozgat Bozok University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Seher\",\"middleName\":\"\",\"lastName\":\"YILMAZ\",\"suffix\":\"\"},{\"id\":531190106,\"identity\":\"971f851a-7ac7-41cb-b4f2-e0b6ae1be606\",\"order_by\":2,\"name\":\"Aslı OKAN\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Yozgat Bozok University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Aslı\",\"middleName\":\"\",\"lastName\":\"OKAN\",\"suffix\":\"\"},{\"id\":531190107,\"identity\":\"92d82889-0b20-4a28-8504-5e08e2558758\",\"order_by\":3,\"name\":\"Sümeyye UÇAR\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Erciyes University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Sümeyye\",\"middleName\":\"\",\"lastName\":\"UÇAR\",\"suffix\":\"\"},{\"id\":531190109,\"identity\":\"6e57704a-d0fb-47ef-801c-588b44ef8cfd\",\"order_by\":4,\"name\":\"Ece EROĞLU\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Yozgat Bozok University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Ece\",\"middleName\":\"\",\"lastName\":\"EROĞLU\",\"suffix\":\"\"},{\"id\":531190111,\"identity\":\"7e62369c-9a89-4905-b875-7c438f87270b\",\"order_by\":5,\"name\":\"Şükrü ATEŞ\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Yozgat Bozok University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Şükrü\",\"middleName\":\"\",\"lastName\":\"ATEŞ\",\"suffix\":\"\"},{\"id\":531190116,\"identity\":\"1f35473c-2042-4e33-89a5-b824a862d939\",\"order_by\":6,\"name\":\"Abdülhadi Cihangir UĞUZ\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"Karamanoglu Mehmetbey University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Abdülhadi\",\"middleName\":\"Cihangir\",\"lastName\":\"UĞUZ\",\"suffix\":\"\"},{\"id\":531190117,\"identity\":\"f8a6e5d6-4f97-405a-b9d6-0e3b3fe1a339\",\"order_by\":7,\"name\":\"Züleyha DOĞANYİĞİT\",\"email\":\"data:image/png;base64,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\",\"orcid\":\"\",\"institution\":\"Yozgat Bozok University\",\"correspondingAuthor\":true,\"prefix\":\"\",\"firstName\":\"Züleyha\",\"middleName\":\"\",\"lastName\":\"DOĞANYİĞİT\",\"suffix\":\"\"}],\"badges\":[],\"createdAt\":\"2025-10-05 16:38:13\",\"currentVersionCode\":1,\"declarations\":\"\",\"doi\":\"10.21203/rs.3.rs-7785868/v1\",\"doiUrl\":\"https://doi.org/10.21203/rs.3.rs-7785868/v1\",\"draftVersion\":[],\"editorialEvents\":[],\"editorialNote\":\"\",\"failedWorkflow\":false,\"files\":[{\"id\":93968156,\"identity\":\"7606c0a0-e4ef-4bb7-ae50-d00bf2ed763a\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:59\",\"extension\":\"docx\",\"order_by\":0,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":3011416,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Manuscript.docx\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/826e45619257cd869e95e7c6.docx\"},{\"id\":93968142,\"identity\":\"519945d9-6150-4ef3-8332-3d5e263eaff1\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"json\",\"order_by\":1,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":9074,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"5dd3746fc5374d09afb7850d6ba902d5.json\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/788c2d0b17ab69a806ed29a5.json\"},{\"id\":93968147,\"identity\":\"47a9ae78-9a0e-415b-b6b7-8de6f04995e4\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"docx\",\"order_by\":2,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":130006,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"EthicalStatement.docx\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/cdfffc0098a348b0ce484e7f.docx\"},{\"id\":93968144,\"identity\":\"2cf28ba0-be72-4b38-b0a0-bfeecb655ff0\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"docx\",\"order_by\":4,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":16817,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"TitlePage.docx\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/0ee56c03811c34f389fa9601.docx\"},{\"id\":93968145,\"identity\":\"69493c06-99da-4671-a399-7ee1f92ce937\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"xml\",\"order_by\":5,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":133483,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"5dd3746fc5374d09afb7850d6ba902d51enriched.xml\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/56687c155218a7634288015d.xml\"},{\"id\":93968152,\"identity\":\"4810d5ea-40df-4f19-a438-c07ef8fa70bb\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"jpeg\",\"order_by\":6,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":703689,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"floatimage1.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/e1c4720ace14d83e86bfee78.jpeg\"},{\"id\":93968157,\"identity\":\"73efbbc8-35fb-46e1-bc83-901467269e8f\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:59\",\"extension\":\"jpeg\",\"order_by\":7,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":2334533,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"floatimage2.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/0b9be199097483a02524508a.jpeg\"},{\"id\":93968465,\"identity\":\"a46175da-125d-4a1c-9137-3b6cec6a9923\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:53:58\",\"extension\":\"png\",\"order_by\":13,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":131885,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage1.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/0fe7daccd6ceba06f29243f7.png\"},{\"id\":93968150,\"identity\":\"19441135-6e9e-45c1-844c-e8d851b5ab14\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"png\",\"order_by\":14,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":803411,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage2.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/fdadd47b1f2375041d7af2c9.png\"},{\"id\":93968163,\"identity\":\"88182a93-c6af-494b-84dd-c87dafa7b220\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:59\",\"extension\":\"png\",\"order_by\":15,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":694613,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage3.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/6ccd1d7dfe2a639b440679e0.png\"},{\"id\":93968151,\"identity\":\"cc8df7ba-b3cf-486b-9479-11f3d58b41fb\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"png\",\"order_by\":16,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":734172,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage4.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/a04bce461f5e97f18e705987.png\"},{\"id\":93968467,\"identity\":\"8e44d98f-833a-4123-9be8-2ff86307e2f2\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:53:59\",\"extension\":\"png\",\"order_by\":17,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":716920,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage5.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/1509ffe5c75f389314046c5c.png\"},{\"id\":93968622,\"identity\":\"c3667c88-1914-45d2-be30-4c7f9eeee023\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 20:01:58\",\"extension\":\"png\",\"order_by\":18,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":746941,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage6.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/e3827700f77a4076a7ce3eee.png\"},{\"id\":93968153,\"identity\":\"170a3c19-affd-4ff8-b688-edd76ab98d98\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"png\",\"order_by\":19,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":733423,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Onlinefloatimage7.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/359e2d5772006f235488c948.png\"},{\"id\":93968161,\"identity\":\"357d46ee-d790-4f8d-a7c7-01d0f91b13cb\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:59\",\"extension\":\"xml\",\"order_by\":20,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":130613,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"5dd3746fc5374d09afb7850d6ba902d51structuring.xml\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/847d72088cd823d505fb0918.xml\"},{\"id\":93968623,\"identity\":\"b42b8e02-8112-476f-ba76-f399959d7e67\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 20:01:59\",\"extension\":\"html\",\"order_by\":21,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"acdc-reference\",\"size\":137994,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"earlyproof.html\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/614538418288e796ea70b96b.html\"},{\"id\":93968141,\"identity\":\"ad1d6d9f-12b8-4fac-82ac-b998e535f141\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"jpeg\",\"order_by\":1,\"title\":\"Figure 1\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":983412,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003e\\u003cstrong\\u003eHematoxylin and eosin staining images of brain tissues of the experimental groups.\\u003c/strong\\u003eBlack arrow indicates neuron degeneration. asterisk indicates vessel dilatation. Magnification is 20X and scale bar is 50 µm.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage2.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/0477b3ce0049c503964cb295.jpeg\"},{\"id\":93968162,\"identity\":\"8f7858b6-5696-4f4e-a772-2a6dd7dcb53a\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:59\",\"extension\":\"jpeg\",\"order_by\":2,\"title\":\"Figure 2\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":1906453,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eA) Immunohistochemistry staining images of TRPV1 in the hippocampus and cortex areas of the experimental groups. Black arrow indicates immunoreactive cells. Magnification 20X and scale bar 50 µm. B) Bar graphs showing the intensity of immunostaining. Two-way analysis of variance, Tukey's and Sidak's multiple comparison tests were applied. Data shown in the bar graph are expressed as mean ± SD. In the female experimental groups, \\u003csup\\u003ea\\u003c/sup\\u003ep\\u0026lt;0.05 indicates a statistically significant difference compared to the control group, \\u003csup\\u003eb\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003ec\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group, \\u003csup\\u003ed\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group. In the male experimental groups, \\u003csup\\u003ee\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the control group, \\u003csup\\u003ef\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003eg\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group and \\u003csup\\u003eh\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage3.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/b44b9960b036d9b2e16a01aa.jpeg\"},{\"id\":93968164,\"identity\":\"f20ec895-9aa7-4cb2-8795-f326c2f9f5ad\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:59\",\"extension\":\"jpeg\",\"order_by\":3,\"title\":\"Figure 3\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":2021994,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eA) Immunohistochemistry staining images of VIP in the hippocampus and cortex areas of the experimental groups. Black arrow indicates immunoreactive cells. Magnification 20X and scale bar 50 µm. B) Bar graphs showing the intensity of immunostaining. Two-way analysis of variance, Tukey's and Sidak's multiple comparison tests were applied. Data shown in the bar graph are expressed as mean ± SD. In the female experimental groups, \\u003csup\\u003ea\\u003c/sup\\u003ep\\u0026lt;0.05 indicates a statistically significant difference compared to the control group, \\u003csup\\u003eb\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003ec\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group, \\u003csup\\u003ed\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group. In the male experimental groups, \\u003csup\\u003ee\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the control group, \\u003csup\\u003ef\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003eg\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group and \\u003csup\\u003eh\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage4.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/93b2e54a7b2e04f5ddeeabb7.jpeg\"},{\"id\":93968143,\"identity\":\"dac9b77c-0f94-4603-8691-dc168751ff46\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"jpeg\",\"order_by\":4,\"title\":\"Figure 4\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":1964857,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eA) Immunohistochemistry staining images of PACAP in the hippocampus and cortex areas of the experimental groups. Black arrow indicates immunoreactive cells. Magnification 20X and scale bar 50 µm. B) Bar graphs showing the intensity of immunostaining. Two-way analysis of variance, Tukey's and Sidak's multiple comparison tests were applied. Data shown in the bar graph are expressed as mean ± SD. In the female experimental groups, \\u003csup\\u003ea\\u003c/sup\\u003ep\\u0026lt;0.05 indicates a statistically significant difference compared to the control group, \\u003csup\\u003eb\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003ec\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group, \\u003csup\\u003ed\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group. In the male experimental groups, \\u003csup\\u003ee\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the control group, \\u003csup\\u003ef\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003eg\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group and \\u003csup\\u003eh\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage5.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/40b71c2c0493c61dadcb0859.jpeg\"},{\"id\":93968148,\"identity\":\"bcd5a461-6e0a-4a15-9d8e-46e880b22a6a\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"jpeg\",\"order_by\":5,\"title\":\"Figure 5\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":2003030,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eA) Immunohistochemistry staining images of P1 in the hippocampus and cortex areas \\u0026nbsp;of the experimental groups. Black arrow indicates immunoreactive cells. Magnification 20X and scale bar 50 µm. B) Bar graphs showing the intensity of immunostaining. Two-way analysis of variance, Tukey's and Sidak's multiple comparison tests were applied. Data shown in the bar graph are expressed as mean ± SD. In the female experimental groups, \\u003csup\\u003ea\\u003c/sup\\u003ep\\u0026lt;0.05 indicates a statistically significant difference compared to the control group, \\u003csup\\u003eb\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003ec\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group, \\u003csup\\u003ed\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group. In the male experimental groups, \\u003csup\\u003ee\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the control group, \\u003csup\\u003ef\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003eg\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group and \\u003csup\\u003eh\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage6.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/3d7a9161bd5c2c509128244b.jpeg\"},{\"id\":93968149,\"identity\":\"48b8242a-5469-48b4-be79-1ad6ca4d5a88\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:58\",\"extension\":\"jpeg\",\"order_by\":6,\"title\":\"Figure 6\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":1999533,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eA) Immunohistochemistry staining images of CGRP in hippocampus and cortex areas of experimental groups. Black arrow indicates immunoreactive cells. Magnification 20X and scale bar 50 µm. B) Bar graphs showing the intensity of immunostaining. Two-way analysis of variance, Tukey's and Sidak's multiple comparison tests were applied. Data shown in the bar graph are expressed as mean ± SD. In the female experimental groups, \\u003csup\\u003ea\\u003c/sup\\u003ep\\u0026lt;0.05 indicates a statistically significant difference compared to the control group, \\u003csup\\u003eb\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003ec\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group, \\u003csup\\u003ed\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group. In the male experimental groups, \\u003csup\\u003ee\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the control group, \\u003csup\\u003ef\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003eg\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group and \\u003csup\\u003eh\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage7.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/366ffb8a9a7e142118572784.jpeg\"},{\"id\":93968927,\"identity\":\"08b2a0f8-d824-46de-8a3b-57c402202cd2\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 20:10:02\",\"extension\":\"pdf\",\"order_by\":0,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"manuscript-pdf\",\"size\":11735545,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"manuscript.pdf\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/332ff226-ee04-4e95-8c47-46a7eb3f754a.pdf\"},{\"id\":93968160,\"identity\":\"53253bac-6641-4a60-87f7-d0931dc06bef\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:45:59\",\"extension\":\"jpg\",\"order_by\":1,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"supplement\",\"size\":125682,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"GraphicalAbstract.jpg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/813b588fcdf88f8861039aa7.jpg\"},{\"id\":93968464,\"identity\":\"b87ddc1f-ce8a-496e-958e-9a1a215af1da\",\"added_by\":\"auto\",\"created_at\":\"2025-10-20 19:53:58\",\"extension\":\"docx\",\"order_by\":2,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"supplement\",\"size\":130006,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"EthicalStatement.docx\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7785868/v1/ecfad667f91a0b8f54787ed7.docx\"}],\"financialInterests\":\"No competing interests reported.\",\"formattedTitle\":\"Effects of Sumatriptan on PACAP, PAC-1, CGRP, VIP, and TRPV-1 Molecules in an Experimental Migraine Model\",\"fulltext\":[{\"header\":\"INTRODUCTION\",\"content\":\"\\u003cp\\u003eMigraine, the second leading cause of disability worldwide, is a common neurological disorder affecting approximately 12% of the population (Stovner et al., \\u003cspan citationid=\\\"CR44\\\" class=\\\"CitationRef\\\"\\u003e2007\\u003c/span\\u003e; Burch et al., \\u003cspan citationid=\\\"CR8\\\" class=\\\"CitationRef\\\"\\u003e2019\\u003c/span\\u003e). Migraine is defined as a recurrent primary headache disorder resulting in attacks lasting 4\\u0026ndash;72 hours. The headache seen in migraine is typically unilateral, episodic, of moderate or severe intensity, and can be aggravated by routine physical activity. Migraine attacks can be associated with nausea, photophobia, and phonophobia (Headache Classification Committee of the International Headache Society, 2013). Despite these data, while the role of the trigeminovascular system in the pathology of migraine is suggested, this condition is not yet fully understood (Ashina et al., \\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e2019\\u003c/span\\u003e; Charles, 2018). In this context, it has been stated that the neurovascular theory induces functional and anatomical changes in the pathophysiology of migraine (Marichal-Cancino et al., \\u003cspan citationid=\\\"CR36\\\" class=\\\"CitationRef\\\"\\u003e2021\\u003c/span\\u003e). In the development of migraine, excessive stimulation of the trigeminovascular system (Hoffman et al., 2019) results in parasympathetic output to the intracranial arteries. This output, in turn, leads to the dilation of intracranial arteries due to the release of signaling molecules such as pituitary adenylate cyclase-activating peptide (PACAP) and calcitonin gene-related peptide (CGRP) (Ashina et al., \\u003cspan citationid=\\\"CR3\\\" class=\\\"CitationRef\\\"\\u003e2017\\u003c/span\\u003e). In this context, targeting signaling molecules like PACAP and CGRP in new approaches could have therapeutic importance. PACAP and its binding receptor, pituitary adenylate cyclase-activating polypeptide type 1 (PAC-1), belong to the vasoactive intestinal peptide (VIP)/secretin/growth hormone-releasing neuropeptide superfamily, which is widely expressed in tissues (Wong et al., \\u003cspan citationid=\\\"CR51\\\" class=\\\"CitationRef\\\"\\u003e1998\\u003c/span\\u003e; Arimura, \\u003cspan citationid=\\\"CR2\\\" class=\\\"CitationRef\\\"\\u003e2007\\u003c/span\\u003e). The existing literature suggests that PACAP and its receptors may play a role in the pathogenesis of migraine-associated headaches (Edvinsson et al., \\u003cspan citationid=\\\"CR13\\\" class=\\\"CitationRef\\\"\\u003e2018\\u003c/span\\u003e). CGRP, a neuropeptide related to signaling pathways in migraine, is produced in peripheral and central neurons and exhibits a potent vasodilatory effect (Brain et al., \\u003cspan citationid=\\\"CR7\\\" class=\\\"CitationRef\\\"\\u003e1985\\u003c/span\\u003e; Rosenfeld et al., \\u003cspan citationid=\\\"CR42\\\" class=\\\"CitationRef\\\"\\u003e1983\\u003c/span\\u003e). In this context, drug trials targeting CGRP in migraine-related attacks have come to the forefront (Tepper, \\u003cspan citationid=\\\"CR48\\\" class=\\\"CitationRef\\\"\\u003e2019\\u003c/span\\u003e). According to the presented data, signaling molecules such as PACAP, PAC-1, VIP, and CGRP stand out as critical molecules in the occurrence of episodic attacks in migraine disease. Additionally, molecules that increase pain sensitivity in migraine-related attacks may also contribute to the enhancement of symptoms. Transient receptor potential (TRP) channels, which are among the molecules that increase pain sensitivity in the perception of migraine-related attacks, consist of six subfamilies: TRPA, TRPC, TRPM, TRPP, TRPL, and TRPV. These channels are primarily expressed on the cell membrane and are part of the cation channel family (Jard\\u0026iacute;n et al., \\u003cspan citationid=\\\"CR30\\\" class=\\\"CitationRef\\\"\\u003e2017\\u003c/span\\u003e). TRP channels contribute to various physiological processes, including temperature regulation and the perception of pain. In this context, transient receptor potential vanilloid 1 (TRPV1) channels, particularly among the TRP channels, are associated with migraine pathogenesis due to the stimulation of meningeal nociceptors related to pain (Benemei and Dussor, \\u003cspan citationid=\\\"CR5\\\" class=\\\"CitationRef\\\"\\u003e2019\\u003c/span\\u003e). Furthermore, the activation of TRP channels can positively affect the release of CGRP from sensory nerve endings (Dussor et al., \\u003cspan citationid=\\\"CR12\\\" class=\\\"CitationRef\\\"\\u003e2014\\u003c/span\\u003e). Therefore, it can be considered that TRPV1, from the TRP channel family, may play a role in the pathophysiological mechanisms related to migraine. In light of all the presented data, targeting PACAP, CGRP, VIP, which cause vasodilation, and TRPV1 channels, which increase pain sensitivity, with potential pharmacological trials in migraine-related attacks may have a positive effect on the acute treatment of migraine. Sumatriptan, one of the drugs used in the acute treatment of migraine, is a 5-HT1 agonist that selectively targets the 5-HT1B and 5-HT1D receptors. Sumatriptan exerts its therapeutic effect in migraine by causing vasoconstriction of dilated meningeal blood vessels, suppressing the release of vasoactive neuropeptides from perivascular trigeminal sensory neurons, and reducing the transmission of pain signals in the trigeminal dorsal horn (Ferrari et al., \\u003cspan citationid=\\\"CR16\\\" class=\\\"CitationRef\\\"\\u003e2002\\u003c/span\\u003e; Goadsby, \\u003cspan citationid=\\\"CR19\\\" class=\\\"CitationRef\\\"\\u003e2007\\u003c/span\\u003e). In another study, the relationship of Sumatriptan with TRPV channels in trigeminal neurons was examined. It was shown that TRPV1 channels are functional in neurons projecting to the cerebral dura and that Sumatriptan suppresses TRPV1 in trigeminal neurons (Evans et al., \\u003cspan citationid=\\\"CR15\\\" class=\\\"CitationRef\\\"\\u003e2012\\u003c/span\\u003e). TRPV1 channels may have a specific role in the pathology of migraine. Additionally, a study conducted in 2021 on 26 migraine patients indicated that early treatment with intravenously administered Sumatriptan prevented PACAP-38-induced migraine (Wienholtz et al., \\u003cspan citationid=\\\"CR50\\\" class=\\\"CitationRef\\\"\\u003e2021\\u003c/span\\u003e). In light of these data, it can be stated that Sumatriptan may affect PACAP-38 in relation to the trigeminovascular system. In the study, the potential effects of Sumatriptan, an agent used in the acute treatment of migraine, on PACAP and its receptor PAC-1, CGRP and VIP, which modulates CGRP, and the TRPV1 channel, which increases pain sensitivity, were investigated, as these have been shown to be related to migraine pathogenesis in the literature. Following this, a treatment group given the Sumatriptan agent was formed, and the animals were dissected. In this context, the immunoreactivity of PACAP, PAC-1, CGRP, VIP, and TRPV1 was evaluated using the immunohistochemistry method.\\u003c/p\\u003e\"},{\"header\":\"MATERIALS AND METHODS\",\"content\":\"\\u003cp\\u003e An experimental migraine animal model was created in the project and approval was received for the study from the Erciyes University Animal Experiments Local Ethics Committee (Ethics committee protocol number: 22/076). In the study, 62 Sprague dawley animals, 31 females and 31 males, weighing 250\\u0026ndash;300 g, were randomly divided into 5 experimental groups and the procedures were observed. During the experiment, rats were housed in cages with a constant ambient temperature (24\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;2\\u0026deg;C), humidity (60\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;5%), and alternating 12-h light and 12-h dark cycles. Experimental groups were designed as follows.\\u003c/p\\u003e\\u003cdiv id=\\\"Sec3\\\" class=\\\"Section2\\\"\\u003e\\u003ch2\\u003eControl group (n\\u0026thinsp;=\\u0026thinsp;10, 5 females, 5 males)\\u003c/h2\\u003e\\u003cp\\u003eRats in this group were fed rodent rearing pellet with free access to food and water for 14 days.\\u003c/p\\u003e\\u003c/div\\u003e\\n\\u003ch3\\u003eSham group (n = 10, 5 females, 5 males)\\u003c/h3\\u003e\\n\\u003cp\\u003eRats were given 1 ml of saline solution (0.9%) intraperitoneally (i.p.) for 14 days.\\u003c/p\\u003e\\n\\u003ch3\\u003eNitroglycerin Migraine group (n = 14, 7 females, 7 males)\\u003c/h3\\u003e\\n\\u003cp\\u003eNitroglycerin agent was administered at 10 mg/kg i.p. to 14 Spraque dawley rats, 7 females and 7 males, in the nitroglycerin-induced migraine group for 14 days. Nitroglycerin agent was dissolved in 30% alcohol, 30% propylene glycol and water, and this solution was diluted in 0.9% saline in a polypropylene tube and injected i.p. into rats (Lai et al., \\u003cspan citationid=\\\"CR34\\\" class=\\\"CitationRef\\\"\\u003e2019\\u003c/span\\u003e).\\u003c/p\\u003e\\n\\u003ch3\\u003eSumatriptan group (n = 14, 7 females, 7 males)\\u003c/h3\\u003e\\n\\u003cp\\u003eRats in this group were given sumatriptan i.p. at a dose of 600 \\u0026micro;g/kg for 14 days (Kurul et al., \\u003cspan citationid=\\\"CR32\\\" class=\\\"CitationRef\\\"\\u003e2008\\u003c/span\\u003e; Casili et al., \\u003cspan citationid=\\\"CR9\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e).\\u003c/p\\u003e\\n\\u003ch3\\u003eNitroglycerin Migraine + Sumatriptan group (n = 14, 7 females, 7 males)\\u003c/h3\\u003e\\n\\u003cp\\u003eRats in this group were given 10 mg/kg i.p. nitroglycerin agent for 14 days. Nitroglycerin agent was dissolved in 30% alcohol, 30% propylene glycol and water. This solution was diluted in 0.9% saline in a polypropylene tube and injected i.p. into rats, and 600 \\u0026micro;g/kg sumatriptan agent was given i.p. 5 minutes after nitroglycerin induction (Lai et al., \\u003cspan citationid=\\\"CR34\\\" class=\\\"CitationRef\\\"\\u003e2019\\u003c/span\\u003e; Kural et al., 2008; Casili et al., \\u003cspan citationid=\\\"CR9\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e).\\u003c/p\\u003e\\u003cp\\u003eWhile the rats were under anesthesia, they were sacrificed with the exsanguination method and the hippocampus and cortex tissue were dissected (Doğanyigit et al., 2023; Hamamcı et al., 2020; Ozkul et al., \\u003cspan citationid=\\\"CR39\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e; Akyuz et al., \\u003cspan citationid=\\\"CR1\\\" class=\\\"CitationRef\\\"\\u003e2021\\u003c/span\\u003e).\\u003c/p\\u003e\\u003cdiv id=\\\"Sec8\\\" class=\\\"Section2\\\"\\u003e\\u003ch2\\u003eHistological Analysis\\u003c/h2\\u003e\\u003cp\\u003eThe tissues were fixed with 10% formaldehyde solution and then washed in tap water (1 night), then dehydration was performed by passing the tissues through increasing alcohol series (70%, 80%, 90% and 100%). Then, the tissue samples were left in xylene (1082984000, Merck) to make them transparent, and after passing through paraffin series, they were embedded in clean paraffin. 5 \\u0026micro;m thick sections taken from paraffin-embedded brain tissue samples using a microtome were stained with Harris hematoxylin and eosin. Histomorphological examination was performed under a light microscope (Olympus BX53) and imaged with a ZEISS Axiscope Colibri 3 digital camera. Neuron degeneration and vascular dilatation in the hippocampus and cortex were evaluated in the brain tissues of the experimental groups. Eosinophilic neuron with pyknotic nuclei, cell swelling or shrinkage was considered as neuron degeneration (Tambe et al., \\u003cspan citationid=\\\"CR47\\\" class=\\\"CitationRef\\\"\\u003e2016\\u003c/span\\u003e; Doğanyiğit et al., \\u003cspan citationid=\\\"CR10\\\" class=\\\"CitationRef\\\"\\u003e2023\\u003c/span\\u003e). Histopathological results in each category were scored as 0\\u0026thinsp;=\\u0026thinsp;none, 1\\u0026thinsp;=\\u0026thinsp;mild, 2\\u0026thinsp;=\\u0026thinsp;moderate and 3\\u0026thinsp;=\\u0026thinsp;severe. Quantification was performed randomly and blindly by two researchers. A minimum of 10 images were used for histomorphological scoring for at least 10 rats (5 females\\u0026thinsp;+\\u0026thinsp;5 males) per experimental group.\\u003c/p\\u003e\\u003c/div\\u003e\\n\\u003ch3\\u003eImmunohistochemistry Analysis\\u003c/h3\\u003e\\n\\u003cp\\u003eAnti-PACAP (Affinity, DF7197), Anti-PAC-1 (Bioss, bs-0198R), Anti-CGRP (Affinity, DF7277), Anti-VIP (Affinity, DF6627) and Anti-TRPV-1 (Cloud-Clone Corp., PAF839Hu01) immunoreactivities were detected by immunohistochemical analysis (Okan et al., \\u003cspan citationid=\\\"CR38\\\" class=\\\"CitationRef\\\"\\u003e2024\\u003c/span\\u003e; Doğanyiğit et al., \\u003cspan citationid=\\\"CR10\\\" class=\\\"CitationRef\\\"\\u003e2023\\u003c/span\\u003e). In summary, after deparaffinization of 5\\u0026micro;m thick sections, citrate buffer was used to open the epitopes (pH: 6.0; Thermo Fischer Scientific, UK, AP-9003-500). The slides were then placed in a 3% hydrogen peroxide solution in methanol to prevent endogenous peroxidase activity. Ultra V block solution (Thermo Fischer Scientific, UK, TA-125-UB) was applied to prevent nonspecific staining. Then, it was incubated with the primary antibody at 4oC overnight. Then, it was incubated with biotinylated goat anti polyvalent secondary antibody (Thermo Fischer Scientific, UK, TP-125-BN) in an oven at 37\\u003csup\\u003eo\\u003c/sup\\u003eC for 40 minutes. After washing several times with PBS, it was incubated with streptavidin peroxidase (Thermo Fischer Scientific, UK, TS-125-HR) in an oven at 37oC for 30 minutes. The antibody complex was visualized by incubation with diaminobenzidine (DAB) chromogen (Thermo Fischer Scientific, UK, TA-125-HD). Then, the sections were counterstained with Gill III Hematoxylin (Merck, Germany, 1.05174.1000). It was dehydrated by passing through an increasing series of alcohol and sealed with a sealer called Entellan. Sections were examined with a ZEISS Axiscope Colibri 3 light microscope. Assessment of immunoreactivity levels was performed with Image J Version 1.46 (National Institutes of Health, Bethesda, Maryland).\\u003c/p\\u003e\\u003cdiv id=\\\"Sec10\\\" class=\\\"Section2\\\"\\u003e\\u003ch2\\u003eStatistical analysis\\u003c/h2\\u003e\\u003cp\\u003eData were expressed as mean\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;SD and analyzed by One-way ANOVA test and Tukey's post-hoc test for parametric tests. In non-parametric tests, independent sample-t test with median minimum-maximum values will be applied. p\\u0026thinsp;\\u0026lt;\\u0026thinsp;0.05 was considered significant in the analyses.\\u003c/p\\u003e\\u003c/div\\u003e\"},{\"header\":\"RESULTS\",\"content\":\"\\u003cdiv id=\\\"Sec12\\\" class=\\\"Section2\\\"\\u003e\\u003ch2\\u003eHistological Findings\\u003c/h2\\u003e\\u003cp\\u003eHistopathological changes in the brain hippocampus and prefrontal cortex areas of the rats in the experimental groups were evaluated under a light microscope after hematoxylin and eosin staining (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig1\\\" class=\\\"InternalRef\\\"\\u003e1\\u003c/span\\u003e). When neuron degeneration in the hippocampus in the nitroglycerin groups was compared, it was observed that it was more severe in males than in females (***p\\u0026thinsp;=\\u0026thinsp;0.0004). While vascular dilation increased in males compared to females in the sumatriptan groups (p\\u0026thinsp;=\\u0026thinsp;0.0045), more dilated blood vessels were observed in females than males in the nitroglycerin groups (p\\u0026thinsp;\\u0026lt;\\u0026thinsp;0.0001). When the cortex was examined, it was found that there was no difference between the experimental groups in terms of neuron degeneration. Vascular dilatation in the cortex was observed to be greater in males than in females in the sumatriptan groups (p\\u0026thinsp;=\\u0026thinsp;0.0223). Neuron degeneration and vascular dilatation in the brain were observed to be significantly increased in the nitroglycerin groups in both male and female groups (Table\\u0026nbsp;1). It was determined that neuron degeneration in the female and male cortices was significantly reduced in the nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups compared to the nitroglycerin groups (Table\\u0026nbsp;1). It was observed that the vascular dilation observed in the hippocampus of nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan females was less severe than that of nitroglycerin females (Table\\u0026nbsp;1).\\u003c/p\\u003e\\u003cp\\u003e\\u003cb\\u003eTable\\u0026nbsp;1.\\u003c/b\\u003e Histopathological damage rates in brain tissues of experimental groups.\\u003c/p\\u003e\\u003ctable border=\\\"1\\\" cellspacing=\\\"0\\\" cellpadding=\\\"0\\\"\\u003e\\n \\u003ctbody\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u0026nbsp;\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd colspan=\\\"2\\\" valign=\\\"top\\\" style=\\\"width: 236px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eHippocampus\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd colspan=\\\"2\\\" valign=\\\"top\\\" style=\\\"width: 208px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eCortex\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eGroups\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 123px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eNeuron Degeneration\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eVascular dilation\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eNeuron Degeneration\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 95px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eVascular dilation\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eControl female\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 123px;\\\"\\u003e\\n \\u003cp\\u003e0.3\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.07\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.32\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.13\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.34\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.08\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 95px;\\\"\\u003e\\n \\u003cp\\u003e0.52\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.19\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eSham female\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 123px;\\\"\\u003e\\n \\u003cp\\u003e0.22\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.08\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.28\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.04\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.34\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.15\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 95px;\\\"\\u003e\\n \\u003cp\\u003e0.5\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.36\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eSumatriptan female\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 123px;\\\"\\u003e\\n \\u003cp\\u003e0.41\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.13\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.3\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.14\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.45\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.19\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 95px;\\\"\\u003e\\n \\u003cp\\u003e0.34\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.16\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eNitroglycerin female\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 123px;\\\"\\u003e\\n \\u003cp\\u003e1.37\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.61\\u003csup\\u003eabc\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e1.77\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.17\\u003csup\\u003eabc\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e2.12\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.14\\u003csup\\u003eabc\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 95px;\\\"\\u003e\\n \\u003cp\\u003e1.55\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.32\\u003csup\\u003eabc\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eNitroglycerin\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003e+Sumatriptan female\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 123px;\\\"\\u003e\\n \\u003cp\\u003e1.55\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.19\\u003csup\\u003eabc\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.78\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.19\\u003csup\\u003eabcd\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e1.52\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.17\\u003csup\\u003eabcd\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 95px;\\\"\\u003e\\n \\u003cp\\u003e0.91\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.31\\u003csup\\u003ecd\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eControl male\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 123px;\\\"\\u003e\\n \\u003cp\\u003e0.18\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.08\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.38\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.30\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.4\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.14\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 95px;\\\"\\u003e\\n \\u003cp\\u003e0.48\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.29\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eSham male\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 123px;\\\"\\u003e\\n \\u003cp\\u003e0.26\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.05\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.28\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.08\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.38\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.13\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 95px;\\\"\\u003e\\n \\u003cp\\u003e0.42\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.13\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eSumatriptan male\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 123px;\\\"\\u003e\\n \\u003cp\\u003e0.54\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.27\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.67\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.38\\u003csup\\u003ef\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.3\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.14\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 95px;\\\"\\u003e\\n \\u003cp\\u003e0.75\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.22\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eNitroglycerin male\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 123px;\\\"\\u003e\\n \\u003cp\\u003e1.98\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.19\\u003csup\\u003eefg\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.88\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.10\\u003csup\\u003eef\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e2.18\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.19\\u003csup\\u003eefg\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 95px;\\\"\\u003e\\n \\u003cp\\u003e1.39\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.21\\u003csup\\u003eefg\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003ctr\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 168px;\\\"\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003eNitroglycerin\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003cp\\u003e\\u003cstrong\\u003e+Sumatriptan male\\u003c/strong\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 123px;\\\"\\u003e\\n \\u003cp\\u003e1.77\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.24\\u003csup\\u003eefg\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e0.61\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.08\\u003csup\\u003ef\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 113px;\\\"\\u003e\\n \\u003cp\\u003e1.7\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.23\\u003csup\\u003eefgh\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003ctd valign=\\\"top\\\" style=\\\"width: 95px;\\\"\\u003e\\n \\u003cp\\u003e1.13\\u003cstrong\\u003e\\u0026plusmn;\\u003c/strong\\u003e0.30\\u003csup\\u003eef\\u003c/sup\\u003e\\u003c/p\\u003e\\n \\u003c/td\\u003e\\n \\u003c/tr\\u003e\\n \\u003c/tbody\\u003e\\n\\u003c/table\\u003e\\u003c/div\\u003e\\u003cdiv id=\\\"Sec13\\\" class=\\\"Section2\\\"\\u003e\\u003ch2\\u003eImmunohistochemistry Findings\\u003c/h2\\u003e\\u003cp\\u003eTRPV1 expression in the hippocampus was lower in nitroglycerin and nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups compared to control, sham and sumatriptan groups. In both nitroglycerin and nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups, TRPV1 expression in the hippocampus was higher in females than in males (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, p\\u0026thinsp;\\u0026lt;\\u0026thinsp;0.0001). TRPV1 expression in the cortex was significantly different between males and females in sham (p\\u0026thinsp;\\u0026lt;\\u0026thinsp;0.0001), sumatriptan (p\\u0026thinsp;=\\u0026thinsp;0.0403) and nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan (p\\u0026thinsp;=\\u0026thinsp;0.0459) groups (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e). In the cortex, TRPV1 expression was decreased in sumatriptan, nitroglycerin and nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups in females and males compared to the control group (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, p\\u0026thinsp;\\u0026lt;\\u0026thinsp;0.05).\\u003c/p\\u003e\\u003cp\\u003e\\u003c/p\\u003e\\u003cp\\u003eWhen the control and sham groups were compared within themselves in terms of VIP expression, a difference was observed between female and male hippocampi. In female hippocampuses, VIP expression decreased in all groups except the control group, whereas in male hippocampuses, VIP expression increased in the nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan group compared to the other groups (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003e). When VIP expression in the cortex was compared between the control, sham and nitroglycerin groups, a difference was observed between male and female rats (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003e). In the cortex, VIP expression was decreased in the other experimental groups compared to the control group in both females and males (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003e).\\u003c/p\\u003e\\u003cp\\u003e\\u003c/p\\u003e\\u003cp\\u003eWhen the control groups were compared within themselves in the hippocampus, a difference was observed between male and female rats in terms of PACAP expression. In Nitroglycerin\\u0026thinsp;+\\u0026thinsp;Sumatriptan groups, increased PACAP expression was observed in both male and female rats in the hippocampus (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig4\\\" class=\\\"InternalRef\\\"\\u003e4\\u003c/span\\u003e). When the control, sham and sumatriptan groups were compared within themselves in the cortex, it was found that there was a difference between male and female rats in terms of PACAP expression. In both male and female rats, PACAP expression in the hippocampus increased in nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups, whereas in the cortex it increased in nitroglycerin groups (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig4\\\" class=\\\"InternalRef\\\"\\u003e4\\u003c/span\\u003e).\\u003c/p\\u003e\\u003cp\\u003e\\u003c/p\\u003e\\u003cp\\u003eIn the hippocampus, when the nitroglycerin group was compared for P1 expression in females and males, it was observed that P1 expression was higher in females than in males. Increased P1 expression was observed in the hippocampus and cortex of female nitroglycerin and nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups. In addition, P1 expression was increased in male hippocampus in nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan group and decreased in nitroglycerin only groups. In the cortex, P1 expression was observed to be different in control, sham, nitroglycerin and nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups in males and females. In male nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups, P1 expression in the cortex was significantly increased (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig5\\\" class=\\\"InternalRef\\\"\\u003e5\\u003c/span\\u003e).\\u003c/p\\u003e\\u003cp\\u003e\\u003c/p\\u003e\\u003cp\\u003eof the experimental groups. Black arrow indicates immunoreactive cells. Magnification 20X and scale bar 50 \\u0026micro;m. B) Bar graphs showing the intensity of immunostaining. Two-way analysis of variance, Tukey's and Sidak's multiple comparison tests were applied. Data shown in the bar graph are expressed as mean\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;SD. In the female experimental groups, \\u003csup\\u003ea\\u003c/sup\\u003ep\\u0026lt;0.05 indicates a statistically significant difference compared to the control group, \\u003csup\\u003eb\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003ec\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group, \\u003csup\\u003ed\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group. In the male experimental groups, \\u003csup\\u003ee\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the control group, \\u003csup\\u003ef\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sham group, \\u003csup\\u003eg\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the sumatriptan group and \\u003csup\\u003eh\\u003c/sup\\u003ep\\u0026lt;0.05 compared to the nitroglycerin group.\\u003c/p\\u003e\\u003cp\\u003eIn the hippocampus, when sumatriptan and nitroglycerin groups were compared within themselves, a difference was observed in terms of CGRP expression in males and females.CGRP expression was significantly increased in the sumatriptan and nitroglycerin sumatriptan groups compared to the control group in males and females (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig6\\\" class=\\\"InternalRef\\\"\\u003e6\\u003c/span\\u003e). When the control, sumatriptan and nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups were compared within themselves in the cortex, a difference was observed in terms of CGRP expression in males and females. Compared to the control group, CGRP expression increased in female sumatriptan groups, while it decreased in males. CGRP expression was similar in female and male nitroglycerin groups. While CGRP expression decreased in female nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan group compared to nitroglycerin group, CGRP expression increased in male nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups compared to nitroglycerin group (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig6\\\" class=\\\"InternalRef\\\"\\u003e6\\u003c/span\\u003e).\\u003c/p\\u003e\\u003cp\\u003e\\u003c/p\\u003e\\u003c/div\\u003e\"},{\"header\":\"DISCUSSION\",\"content\":\"\\u003cp\\u003eIn this study, the effect of sumatriptan on neuropeptides in migraine disease was examined immunoexpressively. Sumatriptan-induced changes in PACAP, PAC-1, CGRP, VIP and TRPV-1 molecules were evaluated immunohistochemically. In both male and female groups, neuronal degeneration and vascular dilatation in the brain were significantly increased in nitroglycerin groups. In addition, sumatriptan significantly decreased neuronal degeneration in the brain in the nitroglycerin groups. In line with the data obtained, it was determined that sumatriptan significantly affected the neuropeptides associated with pain in migraine. Although triptan agents effective in the treatment of acute migraine are well tolerated, the mechanism of action is still debated. In this context, the present study provides an approach that sumatriptan will affect neuropeptides in the brain in migraine disease.\\u003c/p\\u003e\\u003cp\\u003eIn the literature, the relationship between sumatriptan and TRPV1 channels in trigeminal neurons has been examined. It has been shown that TRPV1 channels are functional in neurons projecting to the cerebral dura and sumatriptan suppresses TRPV1 in trigeminal neurons (Evans et al., \\u003cspan citationid=\\\"CR15\\\" class=\\\"CitationRef\\\"\\u003e2012\\u003c/span\\u003e). Our results revealed that TRPV1 immunoactivity in the hippocampus was lower in both the experimental migraine group and the sumatriptan-treated migraine group compared to the control group. These data were significantly higher in the female group compared to males. In the cortex, TRPV1 was significantly decreased in both sexes in the migraine group receiving sumatriptan, the migraine group and the migraine group receiving the sumatriptan agent compared to the control. In line with these data, TRPV1 molecule may play a role in the brain in migraine-induced craniofacial pain. Sumatriptan may be involved in the reduction of migraine-related pain by affecting TRPV channels at the molecular level. Progestogens, estrogens and androgens may play a role in affecting migraine-induced TRPV1 in sumatriptan treatment. TRPV expression may be transcriptionally regulated by sex hormones (Jung et al., \\u003cspan citationid=\\\"CR31\\\" class=\\\"CitationRef\\\"\\u003e2009\\u003c/span\\u003e). Based on these data, the role of sex hormones in the activation, modulation and regulation of the main TRPV channels involved in migraine pathophysiology should be considered.\\u003c/p\\u003e\\u003cp\\u003eParasympathetic neurotransmitters, PACAP38 and VIP, can be released from parasympathetic fibers and activate sensory nerve fibers during migraine attacks. It is known that 5-HT1B/D receptor activation has no effect on circulating VIP and PACAP levels in humans without trigeminovascular activation (Hansen et al., \\u003cspan citationid=\\\"CR26\\\" class=\\\"CitationRef\\\"\\u003e2013\\u003c/span\\u003e). In our study, VIP expression decreased in female hippocampi except for the control group, whereas VIP expression increased in male hippocampi in the nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan group compared to the other groups. In the cortex, VIP expression decreased in the other experimental groups compared to the control group in both females and males. In the literature, it has been shown that 2-hour VIP infusion triggers migraine attacks in migraine patients without aura. VIP was found to have a migraine triggering rate of 71% (Pellesi et al., \\u003cspan citationid=\\\"CR40\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e). In this context, VIP values that differ from the literature should be re-evaluated in the attack pathology of migraine disease in the brain. Given that neuropeptides play an important role in migraine, it may be important to conduct a similar study in the migraine population.\\u003c/p\\u003e\\u003cp\\u003eExperimental studies have shown that PACAP isoforms (PACAP27 and PACAP38) cause migraine attacks after 20 minutes of infusion in migraine patients without aura (Ghanizada et al., \\u003cspan citationid=\\\"CR18\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e). Our results revealed that PACAP immune reactivity was increased in the hippocampus in the sumatriptan-treated migraine group in both females and males. In the cortex, increased PACAP expression was observed in migraine groups of both sexes. Similar to the literature, the relationship between PACAP increase and migraine disease has been shown histopathologically. A study reported that early treatment with intravenously administered sumatriptan prevented migraine caused by PACAP-38 (Wienholtz et al., \\u003cspan citationid=\\\"CR50\\\" class=\\\"CitationRef\\\"\\u003e2021\\u003c/span\\u003e). In 2022, it was reported that sumatriptan had no effect on PACAP38-induced hypersensitivity associated with migraine (Guo et al., \\u003cspan citationid=\\\"CR21\\\" class=\\\"CitationRef\\\"\\u003e2022\\u003c/span\\u003e). In our results, increased PACAP expression was observed in the hippocampus of the migraine experimental group treated with sumatriptan. Therefore, the activity of sumatriptan on PACAP in the brain should be repeated in different groups. Our finding may be interpreted as additional evidence that PACAP-mediated hypersensitivity is different from the known pathways in migraine.\\u003c/p\\u003e\\u003cp\\u003ePreclinical evidence suggests that PACAP38 modulates trigeminal nociceptive activity mainly through PAC1 receptors, while clinical studies report that plasma concentrations of PACAP38 are elevated in cluster headache and migraine spontaneous attacks and return to normal after sumatriptan treatment (Zagami et al., \\u003cspan citationid=\\\"CR52\\\" class=\\\"CitationRef\\\"\\u003e2014\\u003c/span\\u003e). In our study, more PAC1 expression was observed in the hippocampus in females. There is insufficient information on gender-related PAC1 expression in the literature. In this context, PAC1 expression may be modulated in migraine due to gender differences and estrogen dominance. In addition, PAC1 expression increased in the hippocampus and cortex in female nitroglycerin and nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups. P1 expression was found to be significantly increased in the cortex in male nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups. In addition, PAC1 expression was increased in the male hippocampus in the nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan group and decreased in the nitroglycerin only group. These data suggest that the efficacy of sumatriptan may vary in males and females. Our finding draws attention to gender differences in the mechanism of PAC1-mediated PACAP in the known pathways of sumatriptan activity in migraine.\\u003c/p\\u003e\\u003cp\\u003eSumatriptan does not alter cephalic and extrencephalic circulating CGRP in healthy volunteers (Hansen et al., \\u003cspan citationid=\\\"CR27\\\" class=\\\"CitationRef\\\"\\u003e2009\\u003c/span\\u003e). In our study, in the experimental migraine model, CGRP expression was found to be significantly increased in the hippocampus in male and female sumatriptan and nitroglycerin sumatriptan groups compared to the control group. In the cortex, CGRP expression was increased in female sumatriptan groups and decreased in males compared to the control group. In the female nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan group, CGRP expression decreased compared to the nitroglycerin group, while in the male nitroglycerin\\u0026thinsp;+\\u0026thinsp;sumatriptan groups, CGRP expression increased compared to the nitroglycerin group. In this context, the efficacy of sumatriptan on CGRP in migraine may vary depending on the gender parameter. Fluctuations in estrogen levels modulate CGRP receptor signaling in the trigeminovascular system (Labastida-Ram\\u0026iacute;rez et al., \\u003cspan citationid=\\\"CR33\\\" class=\\\"CitationRef\\\"\\u003e2019\\u003c/span\\u003e). The presence of sex hormone receptors in the trigeminovascular system may indicate that trigeminal neurons are sensitive to changes in the levels of these hormones (Gupta et al., \\u003cspan citationid=\\\"CR23\\\" class=\\\"CitationRef\\\"\\u003e2011\\u003c/span\\u003e).\\u003c/p\\u003e\\u003cp\\u003eOur study revealed that sumatriptan, which is used in migraine attacks and belongs to the triptan family, may modulate neuropeptides in the hippocampus and cortex that have been implicated in migraine pathology. Significant sex-related changes in CGRP and TRPV1 immuno expression may be due to endocrine factors such as estrogen hormone. Considering that estrogens modulate the action of nociceptive receptors, female mice may have a lower pain threshold than male mice (Mitrovic et al., \\u003cspan citationid=\\\"CR37\\\" class=\\\"CitationRef\\\"\\u003e2003\\u003c/span\\u003e). However, it may also be important to evaluate the condition during menstruation in females in migraine pathology. The effect of sumatriptan on the menstrual period and migraine attack in females can be discussed in more detail.\\u003c/p\\u003e\\u003cp\\u003eIn pathologies with trigeminal activation such as migraine, 5-HT1B/D receptor activation such as sumatriptan may regulate neuronal VIP and PACAP. The effect of sumatriptan on PACAP- related sensitization in migraine should be evaluated. In our study, increased PACAP expression was observed in the hippocampus of the migraine experimental group treated with sumatriptan. In this context, the significant increase in neuronal PACAP expression may be considered as an effective factor on hypersensitivity. In addition, the mechanism of the PAC1- mediated PACAP-mediated PACAP activity in sumatriptan may also draw attention to gender differences in known pathways in migraine.\\u003c/p\\u003e\\u003cp\\u003eThese data suggest that 5HT1 agonists such as sumatriptan may mediate neuronal neuropeptide changes by regulating the activation of the trigeminal system. This potential therapeutic mechanism of sumatriptan can be evaluated in systems such as migraine or cluster headache to establish its relevance. On the other hand, more detailed analyses are needed to analyze all proteins associated with the trigeminal system, to show which molecules are involved in regulatory mechanisms and to suggest therapeutic approaches for these molecules. Similarly, it is important to reveal the data in this study with different detection methods such as western blot.\\u003c/p\\u003e\"},{\"header\":\"Declarations\",\"content\":\"\\u003cp\\u003e\\u003ch2\\u003eConflict of interest\\u003c/h2\\u003e\\u003cp\\u003eThe author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.\\u003c/p\\u003e\\u003c/p\\u003e\\u003ch2\\u003eFunding\\u003c/h2\\u003e\\u003cp\\u003eThis project was funded by Erciyes University Scientific Research Projects Unit (ERU BAPSIS) (Project support code: FBA-2023-12615).\\u003c/p\\u003e\\u003ch2\\u003eAuthor Contribution\\u003c/h2\\u003e\\u003cp\\u003eFatma \\u0026Ouml;ZT\\u0026Uuml;RK K\\u0026Uuml;P; Validation, Writing \\u0026ndash; original draft. Seher YILMAZ; Formal analysis; Data curation. Aslı OKAN; Formal analysis; Data curation. S\\u0026uuml;meyye U\\u0026Ccedil;AR; Methodology, Conceptualization. Ece EROĞLU; Methodology, Conceptualization. Ş\\u0026uuml;kr\\u0026uuml; ATEŞ; Methodology, Conceptualization. A.Cihangir UĞUZ; Methodology, Conceptualization. Z\\u0026uuml;leyha DOĞANYİĞİT; Writing, review \\u0026amp; editing.\\u003c/p\\u003e\\u003ch2\\u003eAcknowledgement\\u003c/h2\\u003e\\u003cp\\u003eWe thank Erciyes University Scientific Research Projects Unit (ERU BAPSIS) for funding support for this project.\\u003c/p\\u003e\"},{\"header\":\"References\",\"content\":\"\\u003col\\u003e\\u003cli\\u003e\\u003cspan\\u003eAkyuz, E., Doganyigit, Z., Paudel, Y. N., Koklu, B., Kaymak, E., Villa, C., Arulsamy, A., Shaikh, M. F., Devinsky, O. (2021). \\u0026ldquo;Immunoreactivity of Muscarinic Acetylcholine M2 and Serotonin 5-HT2B Receptors, Norepinephrine Transporter and Kir Channels in a Model of Epilepsy\\u0026rdquo;. Life (Basel, Switzerland), 11(4), 276.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eArimura A. (2007). \\u0026ldquo;PACAP: the road to discovery\\u0026rdquo;. Peptides, 28(9), 1617\\u0026ndash;1619.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eAshina, M., Hansen, J. M., \\u0026Aacute; Dunga, B. O., Olesen, J. (2017). \\u0026ldquo;Human models of migraine - short-term pain for long-term gain\\u0026rdquo;. Nature reviews. Neurology, 13(12), 713\\u0026ndash;724.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eAshina, M., Hansen, J. M., Do, T. P., Melo-Carrillo, A., Burstein, R., Moskowitz, M. A. (2019). \\u0026ldquo;Migraine and the trigeminovascular system-40 years and counting\\u0026rdquo;. The Lancet. Neurology, 18(8), 795\\u0026ndash;804.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eBenemei, S., Dussor, G. (2019). \\u0026ldquo;TRP channels and migraine: recent developments and new therapeutic opportunities\\u0026rdquo;. Pharmaceuticals, 12(2), 54.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eBoni, L. J., Ploug, K. B., Olesen, J., Jansen-Olesen, I., Gupta, S. (2009). \\u0026ldquo;The in vivo effect of VIP, PACAP-38 and PACAP-27 and mRNA expression of their receptors in rat middle meningeal artery\\u0026rdquo;. Cephalalgia: an international journal of headache, 29(8), 837\\u0026ndash;847.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eBrain, S. D., Williams, T. J., Tippins, J. R., Morris, H. R., MacIntyre, I. (1985). \\u0026ldquo;Calcitonin gene-related peptide is a potent vasodilator\\u0026rdquo;. Nature, 313(5997), 54\\u0026ndash;56.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eBurch, R. C., Buse, D. C., Lipton, R. B. (2019). \\u0026ldquo;Migraine: Epidemiology, Burden, and Comorbidity\\u0026rdquo;. Neurologic clinics, 37(4), 631\\u0026ndash;649.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eCasili, G., Lanza, M., Filippone, A., Campolo, M., Paterniti, I., Cuzzocrea, S., Esposito, E. (2020). \\u0026ldquo;Dimethyl fumarate alleviates the nitroglycerin (NTG)-induced migraine in mice\\u0026rdquo;. Journal of neuroinflammation, 17(1), 1-Charles A. (2018). \\u0026ldquo;The pathophysiology of migraine: implications for clinical management\\u0026rdquo;. The Lancet. Neurology, 17(2), 174\\u0026ndash;182.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eDoğanyiğit Z, Okan A, Aky\\u0026uuml;z E, Yılmaz S, Ateş Ş, Taheri S, Yılmaz Z, Shaikh MF. (2023). \\u0026ldquo;Can endoplasmic reticulum stress observed in the PTZ-kindling model seizures be prevented with TUDCA and 4-PBA?\\u0026rdquo; Eur J Pharmacol. 2023;960:176072. \\u003cspan class=\\\"ExternalRef\\\"\\u003e\\u003cspan class=\\\"RefSource\\\"\\u003e10.1016/j.ejphar.2023.176072\\u003c/span\\u003e\\u003cspan address=\\\"10.1016/j.ejphar.2023.176072\\\" targettype=\\\"DOI\\\" class=\\\"RefTarget\\\"\\u003e\\u003c/span\\u003e\\u003c/span\\u003e.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eDoğanyiğit, Z., Okan, A., Kaymak, E., Pandır, D., Silici, S. (2020). \\u0026ldquo;Investigation of protective effects of apilarnil against lipopolysaccharide induced liver injury in rats via TLR 4/HMGB- 1/NF-κB pathway\\u0026rdquo;. Biomedicine \\u0026amp; Pharmacotherapy, 125, 109967.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eDussor, G., Yan, J., Xie, J. Y., Ossipov, M. H., Dodick, D. W., Porreca, F. (2014). \\u0026ldquo;Targeting TRP channels for novel migraine therapeutics\\u0026rdquo;. ACS chemical neuroscience, 5(11), 1085\\u0026ndash;1096.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eEdvinsson, L., Tajti, J., Szal\\u0026aacute;rdy, L., V\\u0026eacute;csei, L. (2018). \\u0026ldquo;PACAP and its role in primary headaches\\u0026rdquo;. The journal of headache and pain, 19(1), 1\\u0026ndash;7.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eErdling, A., Sheykhzade, M., Maddahi, A., Bari, F., Edvinsson, L. (2013). \\u0026ldquo;VIP/PACAP receptors in cerebral arteries of rat: characterization, localization and relation to intracellular calcium\\u0026rdquo;. Neuropeptides, 47(2), 85\\u0026ndash;92.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eEvans, M. S., Cheng, X., Jeffry, J. A., Disney, K. E., Premkumar, L. S. (2012). \\u0026ldquo;Sumatriptan inhibits TRPV1 channels in trigeminal neurons\\u0026rdquo;. Headache, 52(5), 773\\u0026ndash;784.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eFerrari, M. D., Goadsby, P. J., Roon, K. I., Lipton, R. B. (2002). \\u0026ldquo;Triptans (serotonin, 5- HT1B/1D agonists) in migraine: detailed results and methods of a meta-analysis of 53 trials\\u0026rdquo;. Cephalalgia, 22(8), 633\\u0026ndash;658.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eGallai, V., Alberti, A., Gallai, B., Coppola, F., Floridi, A., Sarchielli, P. (2003). \\u0026ldquo;Glutamate and nitric oxide pathway in chronic daily headache: evidence from cerebrospinal fluid\\u0026rdquo;. Cephalalgia: an international journal of headache, 23(3), 166\\u0026ndash;174.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eGhanizada, H., Al-Karagholi, M. A., Arngrim, N., Olesen, J., Ashina, M. (2020). \\u0026ldquo;PACAP27 induces migraine-like attacks in migraine patients\\u0026rdquo;. Cephalalgia: an international journal of headache, 40(1), 57\\u0026ndash;67.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eGoadsby, P. J. (2007). \\u0026ldquo;Recent advances in understanding migraine mechanisms, molecules and therapeutics\\u0026rdquo;. Trends in molecular medicine, 13(1), 39\\u0026ndash;44.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eGoadsby, P. J., Edvinsson, L., Ekman, R. (1988). \\u0026ldquo;Release of vasoactive peptides in the extracerebral circulation of humans and the cat during activation of the trigeminovascular system\\u0026rdquo;. Annals of neurology, 23(2), 193\\u0026ndash;196.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eGuo, S., Ernstsen, C., Hay-Schmidt, A., Kristensen, D. M., Ashina, M., Olesen, J., Christensen, S. L. (2022). \\u0026ldquo;PACAP signaling is not involved in GTN-and levcromakalim-induced hypersensitivity in mouse models of migraine\\u0026rdquo;. The Journal of Headache and Pain, 23(1), 155.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eGuo, Z., Czerpaniak, K., Zhang, J., Cao, Y. Q. (2021). \\u0026ldquo;Increase in trigeminal ganglion neurons that respond to both calcitonin gene-related peptide and pituitary adenylate cyclase-activating polypeptide in mouse models of chronic migraine and posttraumatic headache\\u0026rdquo;. Pain, 162(5), 1483\\u0026ndash;1499.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eGupta, S., McCarson, K. E., Welch, K. M. A., Berman, N. E. (2011). \\u0026ldquo;Mechanisms of pain modulation by sex hormones in migraine\\u0026rdquo;. Headache: The Journal of Head and Face Pain, 51(6), 905\\u0026ndash;922.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eHamamci, M., Doganyigit, Z., Silici, S., Okan, A., Kaymak, E., Yilmaz, S., Tokpinar, A., Inan, L. E. (2020). \\u0026ldquo;Apilarnil: A Novel Neuroprotective Candidate\\u0026rdquo;. Acta neurologica Taiwanica, 29(2), 33\\u0026ndash;45.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eHanci, F., Kilinc, Y. B., Kilinc, E., Turay, S., Dilek, M., Kabakus, N. (2021). \\u0026ldquo;Plasma levels of vasoactive neuropeptides in pediatric patients with migraine during attack and attack-free periods\\u0026rdquo;. Cephalalgia: an international journal of headache, 41(2), 166\\u0026ndash;175.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eHansen, J. M., Fahrenkrug, J., Petersen, J., Wienecke, T., Olsen, K. S., Ashina, M. (2013). \\u0026ldquo;Vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) in the circulation after sumatriptan\\u0026rdquo;. Scandinavian journal of pain, 4(4), 211\\u0026ndash;216.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eHansen, J. M., Petersen, J., Wienecke, T., Olsen, K. S., Jensen, L. T., Ashina, M. (2009). \\u0026ldquo;Sumatriptan does not change calcitonin gene-related peptide in the cephalic and extracephalic circulation in healthy volunteers\\u0026rdquo;. The Journal of headache and pain, 10, 85\\u0026ndash;91.).\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eHeadache Classification Committee of the International Headache Society (IHS) (2013). \\u0026ldquo;The International Classification of Headache Disorders, 3rd edition (beta version)\\u0026rdquo;. Cephalalgia: an international journal of headache, 33(9), 629\\u0026ndash;808.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eHoffmann, J., Baca, S. M., Akerman, S. (2019). \\u0026ldquo;Neurovascular mechanisms of migraine and cluster headache\\u0026rdquo;. Journal of cerebral blood flow and metabolism: official journal of the International Society of Cerebral Blood Flow and Metabolism, 39(4), 573\\u0026ndash;594.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eJard\\u0026iacute;n, I., L\\u0026oacute;pez, J. J., Diez, R., S\\u0026aacute;nchez-Collado, J., Cantonero, C., Albarr\\u0026aacute;n, L., Woodard, G. E., Redondo, P. C., Salido, G. M., Smani, T., Rosado, J. A. (2017). \\u0026ldquo;TRPs in Pain Sensation\\u0026rdquo;. Frontiers in physiology, 8, 392.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eJung, C., Fandos, C., Lorenzo, I. M., Plata, C., Fernandes, J., Gen\\u0026eacute;, G. G., Valverde, M. A. (2009). \\u0026ldquo;The progesterone receptor regulates the expression of TRPV4 channel\\u0026rdquo;. Pfl\\u0026uuml;gers Archiv-European Journal of Physiology, 459, 105\\u0026ndash;113.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eKurul, S. H., Demirpence, S., Kiray, M., Tugyan, K., Yilmaz, O., Kose, G. (2008). \\u0026ldquo;Investigation of the immunoreactivities of NOS enzymes and the effect of sumatriptan in adolescent rats using an experimental model of migraine\\u0026rdquo;. The journal of headache and pain, 9(5), 317\\u0026ndash;323.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eLabastida-Ram\\u0026iacute;rez, A., Rubio-Beltr\\u0026aacute;n, E., Villal\\u0026oacute;n, C. M., MaassenVanDenBrink, A. (2019). \\u0026ldquo;Gender aspects of CGRP in migraine\\u0026rdquo;. Cephalalgia, 39(3), 435\\u0026ndash;444.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eLai, T., Chen, L., Chen, X., He, J., Lv, P., Ge, H. (2019). \\u0026ldquo;Rhynchophylline attenuates migraine in trigeminal nucleus caudalis in nitroglycerin-induced rat model by inhibiting MAPK/NF-кB signaling\\u0026rdquo;. Molecular and cellular biochemistry, 461(1\\u0026ndash;2), 205\\u0026ndash;212.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eLiu, J., Wang, G., Dan, Y., Liu, X. (2022). \\u0026ldquo;CGRP and PACAP-38 play an important role in diagnosing pediatric migraine\\u0026rdquo;. The journal of headache and pain, 23(1), 68.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eMarichal-Cancino, B. A., Gonz\\u0026aacute;lez-Hern\\u0026aacute;ndez, A., Guerrero-Alba, R., Medina-Santill\\u0026aacute;n, R., Villal\\u0026oacute;n, C. M. (2021). \\u0026ldquo;A critical review of the neurovascular nature of migraine and the main mechanisms of action of prophylactic antimigraine medications\\u0026rdquo;. Expert review of neurotherapeutics, 21(9), 1035\\u0026ndash;1050.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eMitrovic, I., Margeta-Mitrovic, M., Bader, S., Stoffel, M., Jan, L. Y., Basbaum, A. I. (2003). \\u0026ldquo;Contribution of GIRK2-mediated postsynaptic signaling to opiate and alpha 2-adrenergic analgesia and analgesic sex differences\\u0026rdquo;. Proceedings of the National Academy of Sciences of the United States of America, 100(1), 271\\u0026ndash;276.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eOkan, A., Demir, N., \\u0026amp; Doğanyiğit, Z. (2024). \\u0026ldquo;Linagliptin in combination with ins\\u0026uuml;lin suppresses apoptotic unfolded protein response in ovaries exposed to type 1 diabetes\\u0026rdquo;. Cell biochemistry and function, 42(2), e3898.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eOzkul, Y., Taheri, S., Bayram, K. K., Sener, E. F., Mehmetbeyoglu, E., \\u0026Ouml;ztop, D. B., Aybuga, F., Tufan, E., Bayram, A., Dolu, N., Zararsiz, G., Kianmehr, L., Beyaz, F., Doganyigit, Z., Cuzin, F., Rassoulzadegan, M. (2020). \\u0026ldquo;A heritable profile of six miRNAs in autistic patients and mouse models\\u0026rdquo;. Scientific reports, 10(1), 9011.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003ePellesi, L., Al-Karagholi, M. A., Chaudhry, B. A., Lopez, C. L., Snellman, J., Hannibal, J., Amin, F. M., Ashina, M. (2020). \\u0026ldquo;Two-hour infusion of vasoactive intestinal polypeptide induces delayed headache and extracranial vasodilation in healthy volunteers\\u0026rdquo;. Cephalalgia: an international journal of headache, 40(11), 1212\\u0026ndash;1223.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eP\\u0026eacute;rez-Pereda, S., Toriello-Su\\u0026aacute;rez, M., Ocejo-Vinyals, G., Guiral-Foz, S., Castillo-Obeso, J., Montes-G\\u0026oacute;mez, S., Mart\\u0026iacute;nez-Nieto, R. M., Iglesias, F., Gonz\\u0026aacute;lez-Quintanilla, V., Oterino, A. (2020). \\u0026ldquo;Serum CGRP, VIP, and PACAP usefulness in migraine: a case-control study in chronic migraine patients in real clinical practice\\u0026rdquo;. Molecular biology reports, 47(9), 7125\\u0026ndash;7138.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eRosenfeld, M. G., Mermod, J. J., Amara, S. G., Swanson, L. W., Sawchenko, P. E., Rivier, J., Vale, W. W., Evans, R. M. (1983). \\u0026ldquo;Production of a novel neuropeptide encoded by the calcitonin gene via tissue-specific RNA processing\\u0026rdquo;. Nature, 304(5922), 129\\u0026ndash;135.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eShibata, M., Tang, C. (2021). \\u0026ldquo;Implications of transient receptor potential cation channels in migraine pathophysiology\\u0026rdquo;. Neuroscience bulletin, 37(1), 103\\u0026ndash;116.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eStovner, L. j., Hagen, K., Jensen, R., Katsarava, Z., Lipton, R., Scher, A., Steiner, T., Zwart, J. A. (2007). \\u0026ldquo;The global burden of headache: a documentation of headache prevalence and disability worldwide\\u0026rdquo;. Cephalalgia: an international journal of headache, 27(3), 193\\u0026ndash;210.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eSyed, A. U., Koide, M., Braas, K. M., May, V., Wellman, G. C. (2012). \\u0026ldquo;Pituitary adenylate cyclase-activating polypeptide (PACAP) potently dilates middle meningeal arteries: implications for migraine\\u0026rdquo;. Journal of molecular neuroscience: MN, 48(3), 574\\u0026ndash;583.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eTak\\u0026aacute;cs-Lov\\u0026aacute;sz, K., Kun, J., Acz\\u0026eacute;l, T., Urb\\u0026aacute;n, P., Gyenesei, A., B\\u0026ouml;lcskei, K., Szőke, \\u0026Eacute;., Helyes, Z. (2022). \\u0026ldquo;PACAP-38 Induces Transcriptomic Changes in Rat Trigeminal Ganglion Cells Related to Neuroinflammation and Altered Mitochondrial Function Presumably via PAC1/VPAC2 Receptor-Independent Mechanism\\u0026rdquo;. International journal of molecular sciences, 23(4), 2120.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eTambe, R., Jain, P., Patil, S., Ghumatkar, P., Sathaye, S. (2016). \\u0026ldquo;Antiepileptogenic effects of borneol in pentylenetetrazole-induced kindling in mice\\u0026rdquo;. Naunyn-Schmiedeberg's archives of pharmacology, 389(5), 467\\u0026ndash;475.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eTepper S. J. (2019). \\u0026ldquo;CGRP and headache: a brief review\\u0026rdquo;. Neurological sciences: official journal of the Italian Neurological Society and of the Italian Society of Clinical Neurophysiology, 40 (Suppl 1), 99\\u0026ndash;105.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eTogha, M., Ghorbani, Z., Ramazi, S., Zavvari, F., Karimzadeh, F. (2021). \\u0026ldquo;Evaluation of Serum Levels of Transient Receptor Potential Cation Channel Subfamily V Member 1, Vasoactive Intestinal Polypeptide, and Pituitary Adenylate Cyclase-Activating Polypeptide in Chronic and Episodic Migraine: The Possible Role in Migraine Transformation\\u0026rdquo;. Frontiers in neurology, 12, 770980.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eWienholtz, N., Christensen, C. E., Zhang, D. G., Coskun, H., Ghanizada, H., Al-Karagholi, M. A., Hannibal, J., Egeberg, A., Thyssen, J. P., Ashina, M. (2021). \\u0026ldquo;Early treatment with an international journal of headache, 41(6), 731\\u0026ndash;748.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eWong, A. O., Leung, M. Y., Shea, W. L., Tse, L. Y., Chang, J. P., Chow, B. K. (1998). \\u0026ldquo;Hypophysiotropic action of pituitary adenylate cyclase-activating polypeptide (PACAP) in the goldfish: immunohistochemical demonstration of PACAP in the pituitary, PACAP stimulation of growth hormone release from pituitary cells, and molecular cloning of pituitary type I PACAP receptor\\u0026rdquo;. Endocrinology, 139(8), 3465\\u0026ndash;3479.\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eZagami, A. S., Edvinsson, L., Goadsby, P. J. (2014). \\u0026ldquo;Pituitary adenylate cyclase activating polypeptide and migraine\\u0026rdquo;. Annals of clinical and translational neurology, 1(12), 1036\\u0026ndash;1040.\\u003c/span\\u003e\\u003c/li\\u003e\\u003c/ol\\u003e\"}],\"fulltextSource\":\"\",\"fullText\":\"\",\"funders\":[],\"hasAdminPriorityOnWorkflow\":false,\"hasManuscriptDocX\":true,\"hasOptedInToPreprint\":true,\"hasPassedJournalQc\":\"\",\"hasAnyPriority\":false,\"hideJournal\":false,\"highlight\":\"\",\"institution\":\"\",\"isAcceptedByJournal\":false,\"isAuthorSuppliedPdf\":false,\"isDeskRejected\":\"\",\"isHiddenFromSearch\":false,\"isInQc\":false,\"isInWorkflow\":false,\"isPdf\":false,\"isPdfUpToDate\":true,\"isWithdrawnOrRetracted\":false,\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"identity\":\"bratislava-medical-journal\",\"isNatureJournal\":false,\"hasQc\":true,\"allowDirectSubmit\":false,\"externalIdentity\":\"\",\"sideBox\":\"Learn more about [Bratislava Medical Journal](https://link.springer.com/journal/44411)\",\"snPcode\":\"44411\",\"submissionUrl\":\"https://submission.springernature.com/new-submission/44411/3\",\"title\":\"Bratislava Medical Journal\",\"twitterHandle\":\"\",\"acdcEnabled\":true,\"dfaEnabled\":true,\"editorialSystem\":\"stoa\",\"reportingPortfolio\":\"Springer Hybrid\",\"inReviewEnabled\":true,\"inReviewRevisionsEnabled\":false},\"keywords\":\"Migraine, Neurovascular Theory, Nitroglycerin, Sumatriptan, Trigeminal neurons\",\"lastPublishedDoi\":\"10.21203/rs.3.rs-7785868/v1\",\"lastPublishedDoiUrl\":\"https://doi.org/10.21203/rs.3.rs-7785868/v1\",\"license\":{\"name\":\"CC BY 4.0\",\"url\":\"https://creativecommons.org/licenses/by/4.0/\"},\"manuscriptAbstract\":\"\\u003cp\\u003eMigraine is a chronic neurological disorder characterized by severe headache. In the pathology of migraine, it is suggested that increased parasympathetic output to intracranial arteries due to excessive stimulation of the trigeminovascular system is responsible. The release of PACAP, VIP, and CGRP signaling molecules due to excessive stimulation leads to the dilation of intracranial arteries, causing headaches. Sumatriptan exerts its therapeutic effect by causing vasoconstriction of dilated meningeal blood vessels in migraine and suppressing the release of vasoactive neuropeptides from trigeminal sensory neurons. However, the relationship between the Sumatriptan agent and the signaling pathways in migraine pathology has not been sufficiently studied. In this study, we aimed to determine the potential therapeutic efficacy of the Sumatriptan agent on PACAP, PAC-1, CGRP, VIP, and TRPV1 molecules in an experimental migraine model.\\u003c/p\\u003e\\u003cp\\u003eIn the study, 62 Sprague Dawley rats (31 males and 31 females) weighing 250\\u0026ndash;300 grams were randomly divided into five experimental groups: Control, Sham, Migraine, Sumatriptan, and Migraine-Sumatriptan, with procedures followed accordingly. At the end of the study, histopathological evaluation of brain tissues taken from the animals was performed, and the immunoreactivity of PACAP, PAC-1, CGRP, VIP, and TRPV1 proteins was assesse immunohistochemically.\\u003c/p\\u003e\\u003cp\\u003eAs a result, it was observed that the histopathological findings of the sumatriptan treatment group were reduced compared to the migraine groups. Based on the obtained data, it was determined that sumatriptan significantly affected the neuropeptides associated with pain in migraine. However, more detailed analyses are still needed in this regard.\\u003c/p\\u003e\",\"manuscriptTitle\":\"Effects of Sumatriptan on PACAP, PAC-1, CGRP, VIP, and TRPV-1 Molecules in an Experimental Migraine Model\",\"msid\":\"\",\"msnumber\":\"\",\"nonDraftVersions\":[{\"code\":1,\"date\":\"2025-10-20 19:45:53\",\"doi\":\"10.21203/rs.3.rs-7785868/v1\",\"editorialEvents\":[{\"type\":\"communityComments\",\"content\":0},{\"type\":\"decision\",\"content\":\"Revision requested\",\"date\":\"2025-11-27T12:15:14+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"editorInvitedReview\",\"content\":\"\",\"date\":\"2025-11-25T15:36:27+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"editorInvitedReview\",\"content\":\"\",\"date\":\"2025-11-11T16:59:23+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"94194886089423783968198515228887870887\",\"date\":\"2025-10-29T19:42:17+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"47284166482571949263468039446508579051\",\"date\":\"2025-10-22T21:22:45+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"109210535757484593935575313412437052080\",\"date\":\"2025-10-22T14:40:18+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"7288002985088419492297003320988877416\",\"date\":\"2025-10-22T13:51:40+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"315575633729229055183576880155818423740\",\"date\":\"2025-10-20T23:19:36+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"106049421041317821406784863603236163339\",\"date\":\"2025-10-07T11:41:10+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewersInvited\",\"content\":\"\",\"date\":\"2025-10-07T11:31:26+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"editorAssigned\",\"content\":\"\",\"date\":\"2025-10-06T10:31:36+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"checksComplete\",\"content\":\"\",\"date\":\"2025-10-06T10:30:44+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"submitted\",\"content\":\"Bratislava Medical Journal\",\"date\":\"2025-10-05T16:23:50+00:00\",\"index\":\"\",\"fulltext\":\"\"}],\"status\":\"published\",\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"identity\":\"bratislava-medical-journal\",\"isNatureJournal\":false,\"hasQc\":true,\"allowDirectSubmit\":false,\"externalIdentity\":\"\",\"sideBox\":\"Learn more about [Bratislava Medical Journal](https://link.springer.com/journal/44411)\",\"snPcode\":\"44411\",\"submissionUrl\":\"https://submission.springernature.com/new-submission/44411/3\",\"title\":\"Bratislava Medical Journal\",\"twitterHandle\":\"\",\"acdcEnabled\":true,\"dfaEnabled\":true,\"editorialSystem\":\"stoa\",\"reportingPortfolio\":\"Springer Hybrid\",\"inReviewEnabled\":true,\"inReviewRevisionsEnabled\":false}}],\"origin\":\"\",\"ownerIdentity\":\"de0dbb13-28f0-490e-aa87-457aac8974c6\",\"owner\":[],\"postedDate\":\"October 20th, 2025\",\"published\":true,\"recentEditorialEvents\":[],\"rejectedJournal\":[],\"revision\":\"\",\"amendment\":\"\",\"status\":\"under-review\",\"subjectAreas\":[],\"tags\":[],\"updatedAt\":\"2025-12-10T09:23:36+00:00\",\"versionOfRecord\":[],\"versionCreatedAt\":\"2025-10-20 19:45:53\",\"video\":\"\",\"vorDoi\":\"\",\"vorDoiUrl\":\"\",\"workflowStages\":[]},\"version\":\"v1\",\"identity\":\"rs-7785868\",\"journalConfig\":\"researchsquare\"},\"__N_SSP\":true},\"page\":\"/article/[identity]/[[...version]]\",\"query\":{\"redirect\":\"/article/rs-7785868\",\"identity\":\"rs-7785868\",\"version\":[\"v1\"]},\"buildId\":\"XKTyCvWXoU3ODBz1xrDgd\",\"isFallback\":false,\"isExperimentalCompile\":false,\"dynamicIds\":[84888],\"gssp\":true,\"scriptLoader\":[]}","source_license":"CC-BY-4.0","license_restricted":false}