Activation of the Transcription Factor NF-κB in the Nucleus Trigeminalis Caudalis in an Animal Model of Migraine

doi:10.1016/j.neuro.2005.02.005

NeuroToxicology

Volume 26, Issue 5, October 2005, Pages 795-800

https://doi.org/10.1016/j.neuro.2005.02.005 Get rights and content

Abstract

The infusion of nitroglycerin (NTG) induces an inflammatory state in perivascular meningeal tissues of rat via the activation, inter alia, of nuclear factor kappa B (NF-κB). This phenomenon has been related to the mechanisms involved in the pathophysiology of migraine, a common neurovascular disorder. In the present study, we sought to elucidate whether NF-κB activation might have a role in the determinism of migraine attacks also at the neuronal level. Therefore, we investigated the transcriptional activity of NF-κB in the brainstem of rats systemically injected with NTG and killed 4 h later. Activation of NF-κB in brain areas was detected by means of both the immunohistochemical technique and the Western blot analysis. A significant increase of nuclear immunostaining of p65, an indicator of NF-κB activation, was detected in lamina I and II of nucleus trigeminalis caudalis in rats injected with NTG when compared with the control group. Western blot analysis confirmed the activation of the NF-κB pathway showing an increase in the optical density of p65 in nuclear extracts of lower brainstem of rats injected with the nitric oxide (NO) donor. The present study contributes to expand on our understanding of the complex mechanisms by which NTG may trigger migraine-like headaches in migraineurs. Furthermore, these findings pave the way to new bio-molecular and pharmacological avenues for the development of innovative migraine therapies.

Section snippets

INTRODUCTION

Migraine is a complex disease that involves the mediation of central and peripheral components of the trigeminal pain pathway. Several lines of evidence suggest that pain in migraine is related to the activation of the trigeminovascular system and to the activation of the nucleus trigeminal caudalis (NTC), which conveys the information from the periphery to higher sites.

Infusion of nitroglycerin (NTG) induces spontaneous-like headaches in migraineurs with a delay of hours, but not in control

Animals

Male Sprague–Dawley rats (weight 180–220 g) were divided into groups of four to six animals. All steps were taken at every stage of the experiments to avoid suffering to the animals. The rats were housed in plastic boxes with water and food available ad libitum and kept on a 12-h light:12-h dark cycle.

In the treated groups, animals were injected intraperitoneally with NTG (10 mg/kg b.w.) and perfused transcardially 4 h later (previously identified as the time of maximal Fos expression) with saline

RESULTS

Data regarding Fos expression following the administration of NTG confirmed our previous studies: NTG induced a significant activation of neurons located in lamina I and II of NTC (Fig. 1). The number of p65-immunoreactive neurons in this nucleus was significantly higher in the group treated with NTG when compared with the control group (Fig. 2). The highest concentration of p65-positive nuclei was observed in lamina I, and to a lesser extent in lamina II. Western blotting analysis confirmed

DISCUSSION

In the neuroscience and neurological fields, a general agreement exists on the trigeminovascular mediation of migraine attacks. Actually trigeminovascular-mediated neurogenic inflammation in perivascular tissues within the dura mater has been widely accepted as the “efferent arm” of the process (Moskowitz, 1992). NO and cytokines may contribute to inflammation and sensitization of primary afferents of the trigeminovascular system (Opree and Kress, 2000) and some authors have suggested that the

ACKNOWLEDGMENT

This study was supported by a grant of the Ministry of Health (RC 2002-2004) and by EU Eurohead, LSHM-CT-2004-504837.

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Network pharmacology and experimental verification to explore the anti-migraine mechanism of Yufeng Ningxin Tablet
2023, Journal of Ethnopharmacology
Yufeng Ningxin Tablet (YNT) is a traditional Chinese medicine formula, that has been used clinically to treat migraine for many years. It is composed of one herb Pueraria lobata var. lobata (Willd.) Ohwi (Relevant Chinese name: Gegen). Previously, it has been recorded by traditional Chinese doctor that Gegen could be used as medicine to treat migraine. However, the underlying mechanism of action remains to be investigated.
It was to explore the effect and mechanism of YNT on migraine based on network pharmacology and experimental verification.
First, with the network pharmacology, the effective chemical components and target genes of YNT were filtrated, the YNT-compound-migraine-targets network was constructed. The protein-protein interaction network (PPI) and literature reports were combined to identify potential targets of YNT in the treatment of migraine. Then, the representative compounds of YNT were characterized by LC-MS/MS and the major effect components were identified. Finally, the prediction results of network pharmacology were verified by animal and cell experiments.
7 bioactive components of YNT could act on 97 migraine potential targets. The 5 bioactive components could be characterized comprehensively of YNT. The key therapeutic targets and pathways were collected including 5-HT, CGRP, inflammation and nociceptive factors, and NF-κB signaling pathway. Animal experiments showed that YNT could increase the expression level of 5-HT and reduce the expression of CGRP, NF-κB, c-fos and IL-1β. YNT could inhibit LPS-induced neuroinflammation by NF-κB in BV2 cells in vitro. Western blotting analysis results showed YNT inhibited the NF-κB and phospho–NF–κB levels.
It is the first time to verify the consistency between the metabolic components of YNT by LC-MS/MS and the active components predicted by network pharmacology. Meanwhile, the potential mechanism of YNT in the treatment of migraine was studied by combining network pharmacology and in vitro and in vivo experiments.
Disulfiram and metformin combination anticancer effect reversible partly by antioxidant nitroglycerin and completely by NF-κB activator mebendazole in hamster fibrosarcoma
2021, Biomedicine and Pharmacotherapy
Citation Excerpt :
Necroptosis can be prevented by scavenging ROS production in heat stress-induced intestinal injury [101]. Expected NF-κB inhibitory effect of nitroglycerin [48] can be opposite and manifested in NF-κB stimulation, for example in nucleus trigeminalis caudatus of migraine animal model [28]. Also, nitroglycerin can be without any effect on NF-κB [26].
We investigated the anticancer effect of disulfiram and metformin combination on fibrosarcoma in hamsters. Hamsters of both sexes (~ 70 g) were randomly allocated to control and experimental groups (8 animals per group). In all 10 groups, 2 × 10⁶ BHK-21/C13 cells in 1 ml were injected subcutaneously into the animals’ backs. Peroral treatments were carried out with disulfiram 50 mg/kg daily, or with metformin 500 mg/kg daily, or with their combination. Validation and rescue grups were treated by double doses of the single therapy and by the combination with addition of rescue daily doses of ROS inhibitor nitroglycerin 25 mg/kg or NF-κB stimulator mebendazole 460 mg/kg, via a gastric probe after tumor inoculation. After 19 days all animals were sacrificed. Blood samples were collected for hematological and biochemical analyses, the tumors were excised and weighed, and their diameters and volumes were measured. The tumor samples were pathohistologically and immunohistochemically assessed (Ki-67, PCNA, CD34, CD31, COX4, Cytochrome C, GLUT1, iNOS), and the main organs were toxicologically tested. The combination of disulfiram and metformin significantly inhibited fibrosarcoma growth in hamsters without toxicity, compared to monotherapy or control. The single treatments did not show significant antisarcoma effect. Co-treatment with nitroglycerin partly rescued tumor progression, probably by ROS inhibition, while mebendazole completely blocked anticancer activity of the disulfiram and metformin combination, most likely by NF-κB stimulation. Combination of disulfiram with metformin may be used as an effective and safe candidate for novel nontoxic adjuvant and relapse prevention anticancer therapy.
Characterization of the peripheral FAAH inhibitor, URB937, in animal models of acute and chronic migraine
2021, Neurobiology of Disease
Inhibiting the activity of fatty-acid amide hydrolase (FAAH), the enzyme that deactivates the endocannabinoid anandamide, enhances anandamide-mediated signaling and holds promise as a molecular target for the treatment of human pathologies such as anxiety and pain. We have previously shown that the peripherally restricted FAAH inhibitor, URB937, prevents nitroglycerin-induced hyperalgesia – an animal model of migraine - and attenuates the activation of brain areas that are relevant for migraine pain, e.g. trigeminal nucleus caudalis and locus coeruleus. The current study is aimed at profiling the behavioral and biochemical effects of URB937 in animal models of acute and chronic migraine. We evaluated the effects of URB937 in two rat models that capture aspects of acute and chronic migraine, and are based on single or repeated administration of the vasodilating drug, nitroglycerin (NTG). In addition to nocifensive behavior, in trigeminal ganglia and medulla, we measured mRNA levels of neuropeptides and pro-inflammatory cytokines along with tissue levels of anandamide and palmitoylethanolamide (PEA), an endogenous agonist of peroxisome proliferator-activated receptor type-a (PPAR-a), which is also a FAAH substrate. In the acute migraine model, we also investigated the effect of subtype-selective antagonist for cannabinoid receptors 1 and 2 (AM251 and AM630, respectively) on nocifensive behavior and on levels of neuropeptides and pro-inflammatory cytokines. In the acute migraine paradigm, URB937 significantly reduced hyperalgesia in the orofacial formalin test when administered either before or after NTG. This effect was accompanied by an increase in anandamide and PEA levels in target neural tissue, depended upon CB1 receptor activation, and was associated with a decrease in calcitonin gene-related peptide (CGRP), substance P and cytokines TNF-alpha and IL-6 mRNA. Similar effects were observed in the chronic migraine paradigm, where URB937 counteracted NTG-induced trigeminal hyperalgesia and prevented the increase in neuropeptide and cytokine transcription.
The results show that peripheral FAAH inhibition by URB937 effectively reduces both acute and chronic NTG-induced trigeminal hyperalgesia, likely via augmented anandamide-mediated CB1 receptor activation. These effects are associated with inhibition of neuropeptidergic and inflammatory pathways.
FAAH inhibition as a preventive treatment for migraine: A pre-clinical study
2020, Neurobiology of Disease
Citation Excerpt :
The data gathered in this study do not warrant definite conclusions regarding the precise mechanisms that are engaged selectively by the pre-NTG dosing of URB597. One interesting hypothesis is that the simultaneous elevations of both AEA and PEA may be required to prevent the activation of the inflammatory pathway triggered by NTG (Greco et al., 2005). Further experimental data are needed to confirm this speculation.
Fatty-acid amide hydrolase (FAAH) is an intracellular serine hydrolase that catalyzes the cleavage of endogenous fatty-acid amides, including the endocannabinoid anandamide (AEA). We previously reported that the peripherally restricted FAAH inhibitor URB937, which selectively increases AEA levels outside the central nervous system, reduces hyperalgesia and c-Fos expression in the trigeminal nucleus caudalis (TNC) and the locus coeruleus in an animal model of migraine based on nitroglycerin (NTG) administration.
To further investigate the relevance of FAAH inhibition in the NTG animal model of migraine by testing the effects of the globally active FAAH inhibitor URB597.
Our experimental approach involved mapping neuronal c-Fos protein expression, measurement of AEA levels in brain areas and in trigeminal ganglia, evaluation of pain-related behavior and quantification of molecular mediators in rats that received URB597 (2 mg/kg i.p.) either before or after NTG administration (10 mg/kg, i.p.).
Pre-treatment with URB597 significantly reduced c-Fos immunoreactivity in the TNC and inhibited NTG-induced hyperalgesia in the orofacial formalin test. This behavioral response was associated with a decrease in neuronal nitric oxide synthase, calcitonin gene-related peptide and cytokine gene expression levels in central and peripheral structures. Administration of URB597 after NTG had no such effect.
The findings suggest that global FAAH inhibition may offer a therapeutic approach to the prevention, but not the abortive treatment, of migraine attacks. Further studies are needed to elucidate the exact cellular and molecular mechanisms underlying the protective effects of FAAH inhibition.
Nitroglycerin as a comparative experimental model of migraine pain: From animal to human and back
2019, Progress in Neurobiology
Citation Excerpt :
The increased release of artemin from the dural vasculature may play a role in the sensitization of dural afferents (McIlvried et al., 2010) and contribute to meningeal inflammation (Malin et al., 2006; Shang et al., 2016). Systemic NTG administration activates inflammatory pathways (Greco et al., 2005, 2015b; Greco et al., 2016), probably as a consequence of the increased availability of NO in trigeminovascular terminals. NTG injection indeed induced an up-regulation of pro-inflammatory genes, with a subsequent delayed inflammatory reaction in the dura mater and central areas of the rat (Reuter et al., 2002; Kim et al., 2008; Greco et al., 2016).
Migraine is a disease for which there is still no defined pathophysiological etiology and few translational models. The organic nitrate nitroglycerin has been in use as an experimental model of migraine in both human and animal studies for several years. The drug produces a number of effects within the head, that includes blood vessels, nerves and brain areas that may produce a response similar to a migraine attack in predisposed subjects. A better understanding of the nature of these changes and how well they parallel a true migraine attack would allow for a translational model to better understand some of the mechanisms involved in the generation of a migraine attack. The present review summarizes the known body of knowledge of nitroglycerin effects evaluated in humans and animals as it relates to potential mechanisms associated with migraine headaches.
Pharmacodynamic action and mechanism of Du Liang soft capsule, a traditional Chinese medicine capsule, on treating nitroglycerin-induced migraine
2017, Journal of Ethnopharmacology
Du Liang soft capsule (DL) is a traditional Chinese medicine for treating migraines; it is made from two Chinese herbs, including Ligusticum striatum DC., root; Angelica dahurica (Hoffm.) Benth. & Hook.f. ex Franch. & Sav., root.
In the present study, we aimed to elucidate the pharmacodynamic action of DL and its mechanism in an animal model of migraines induced by glyceryl trinitrate (GTN).
Sixty rats were randomly divided into six groups, including a normal control group, model control group, positive group (Sumatriptan 0.006 g kg⁻¹), and three DL groups (0.44, 1.31 and 3.93 g kg⁻¹). All rats were intragastrically treated with the corresponding treatment for 7 consecutive days, and they were subcutaneously injected with GTN (10 mg kg⁻¹) 30 min after the last treatment, except in the normal control group. After model establishment, the behaviors of all rats, including head scratching, cage climbing, and the development of red ears were observed continuously by digital camera every 30 min for 3 h. Four hours after GTN treatment, all rats were anaesthetized and the blood and tissue samples were collected. Plasma calcitonin gene related to peptide (CGRP) and endothelin (ET) levels were measured using the radioimmunoassay method, and serum NO was determined by the colorimetric method. Afterwards, the brainstem tissues were dissected and washed with physiological saline, and divided evenly into two parts. One part was used to test the monoamine levels, including levels of 5-hydroxytryptamine (5-HT), norepinephrine (NE) and dopamine (DA), by the fluorometric method, and the other part was used to determine the nuclear factor kappaB (NF-κB) p65, nuclear c-fos, inducible nitric oxide synthase (iNOS), interleukin (IL)-1β (IL-1β), and cyclooxygenase-2 (COX-2) levels by Western blot analysis.
In the pharmacodynamic action assay, DL (1.31 and 3.93 g kg⁻¹) greatly improved the abnormal behaviors of migraine rats, including head scratching and cage climbing, and the development of red ears. In the mechanism assay, compared with the control group, the plasma CGRP and serum NO levels and the brainstem 5-HT, NE and DA levels in the DL administration groups were significantly decreased; and the plasma ET levels were remarkably increased. Moreover, down-regulation of NF-κB p65, c-fos and pro-inflammatory cytokines, including iNOS, IL-1β and COX-2 in the brainstem in the DL administration groups were observed by Western blot analysis.
The above results suggested that DL has a therapeutic effect on migraines, and its mechanism may be related to adjusting the level of neurotransmitters and vasoactive substances, consequently relieving neurogenic inflammation.

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Activation of the Transcription Factor NF-κB in the Nucleus Trigeminalis Caudalis in an Animal Model of Migraine

Abstract

Section snippets

INTRODUCTION

Animals

RESULTS

DISCUSSION

ACKNOWLEDGMENT

Eur J Pharmacol

Brain Res

Trends Pharmacol Sci

Neuropharmacology

J Chem Neuroanat

J Biol Chem

Nuclear factor-κB

Int J Biochem Cell Biol

Release of substance P, calcitonin gene-related peptide and prostaglandin E2 from dura mater encephali following electrical chemical stimulation in vitro

Neuroscience

Biphasic response to nitric oxide of spinal trigeminal neurons with meningeal input in rat—possible implications for the pathophysiology of headaches

J Neurophysiol

Spinal NF-kB activation induces COX-2 upregulation and contributes to inflammatory pain hypersensitivity

Eur J Neurosci

Nitric oxide supersensitivity: a possible molecular mechanism of migraine pain

Neuroreport

Involvement of the proinflammatory cytokines tumor necrosis factor-a, IL-1B, and IL-6 but IL-8 in the development of heat hyperalgesia: effects on heat-evoked calcitonin gene-related peptide release from rat skin

J Neurosci

Systemic nitroglycerin increases nNOS levels in rat trigeminal nucleus caudalis

Neuroreport