Migraine Research is a Headache: Evaluating the Role of Dural Complete Freund's Adjuvant-Induced Inflammation and Transient Receptor Potential Melastatin 3 in Preclinical Models Used in Migraine Research

Background: Headaches have burdened humanity for centuries, with migraine, a primary headache disor-der, remaining a leading cause of disability worldwide, affecting nearly one-third of the global population. Characterized by recurrent, long-lasting, pulsating unilateral headaches, the pathophysiology of migraine is still not fully understood despite recent research advances. The neuropeptide calcitonin gene-related peptide (CGRP) plays an important role in migraine and is a key target within the trigeminovascular sys-tem (TGVS), with therapies alleviating symptoms in about half of patients. However, more insights into the mechanisms of CGRP in the TGVS are needed for improved treatments. Neurogenic inflammation and transient receptor potential (TRP) channels have also been implicated in CGRP signaling pathways. How-ever, their precise roles in migraine pathophysiology remain uncertain. Preclinical models have been in-strumental in advancing our understanding of migraine. For instance, inducing meningeal inflammation using inflammatory mediators, such as complete Freund’s adjuvant (CFA), has shown promise as a poten-tial migraine model. However, behavioral data, a critical aspect of model validation, has not been ac-quired.
Aims: In Study I, we aimed to investigate the effect of dural CFA on behavioral outcomes and assess whether known migraine treatments, including the monoclonal CGRP antibody fremanezumab and onabotulinumtoxinA (BoNT/A), could reverse these effects. Additionally, we sought to evaluate the utility of the dural CFA model as a preclinical migraine model. In Study II, we aimed at investigating the TRP Melastatin 3 (TRPM3) channel, which has demonstrated relevance in CGRP-related pathways outside mi-graine contexts. However, its role in migraine-relevant tissues and behaviors was unexplored. We aimed to investigate the effects of a TRPM3 agonist, CIM0216, on CGRP release within the TGVS, its vasomotor effects on meningeal blood vessels, its localization within CGRP-related tissues, and its impact on perior-bital sensitivity. Additionally, we wanted to examine potential sex differences and evaluated the broader role of TRPM3 in migraine research as a potential therapeutic target.
Results: In Paper I, when we induced dural CFA it had no significant impact on behavioral outcomes in the open field or light/dark box tests. However, when we did the electronical von Frey test it lowered withdrawal thresholds in the periorbital region, indicating cephalic hypersensitivity, while we did not see any effects in the plantar region. We did not see alleviated periorbital hypersensitivity with the treatment with fremanezumab.
In Paper II, after we induced dural CFA, we observed unilateral periorbital mechanical hypersensitivity, most pronounced at the administration site. We did not see a reversal effect of the hypersensitivity with the treatment with BoNT/A. We observed a reduction of baseline CGRP release in the more inflamed hemiskull and inhibited capsaicin-stimulated CGRP release in the less inflamed hemiskull, in the BoNT/A treated rats. We observed the presence of cleaved synaptosomal-associated protein-25, indicating BoNT/A activity, in the trigeminal ganglia (TG) of BoNT/A treated rats.
In Paper III we compiled knowledge and data from various studies investigating the effects of inflamma-tion in preclinical models used in migraine research. This included models of CFA-induced inflammation. In the review we examined the validity of these models in the context of migraine research and explored whether inflammation should be considered a part of migraine pathophysiology.
In Paper IV, when we applied CIM0216, a TRPM3 agonist, it triggered CGRP release at 100 µM concen-trations in the TG and dura mater of both male and female rats. We observed an inhibition of CGRP re-lease by the TRPM3 antagonist isosakuranetin in the TG but not in the dura mater. Furthermore, we ob-served that CIM0216 also induced vasodilation in the middle cerebral artery and middle meningeal artery, though this effect was not blocked by isosakuranetin or fremanezumab. We noticed TRPM3 localization in small- to medium-sized TG neurons, nerve fibers in the dura mater, and vascular smooth muscle cells in meningeal blood vessels. Finally, we observed no apparent sex differences, besides potential higher CGRP release in females.
Discussion:
Dural CFA induced unilateral periorbital hypersensitivity but did not affect other behavioral outcomes. Furthermore, known migraine treatments, such as fremanezumab and BoNT/A, were unable to reverse these effects. Based on the insights from Paper III regarding preclinical inflammation models in migraine research, dural CFA demonstrated both significant strengths and notable limitations as a preclinical model for studying migraine. It mimics cephalic ipsilateral allodynia observed in migraine patients but does not mimic other behavioral or treatment-responsive features . This suggests the model could be valuable for studying non-responder populations or mechanisms of chronic sensitization driven by in-flammation, which may help to understand ipsilateral cephalic allodynia in patients. In the TRPM3 study, TRPM3 demonstrated a functional role within the TGVS relevant to migraine, suggesting it could be a potential therapeutic target. However, the lack of significant behavioral effects necessitates further research to elucidate its role in migraine pathophysiology.
Conclusion:
While the dural CFA model contributes to understanding specific aspects of migraine pathophysiology, it cannot be used to fully cover the complexity of migraine. TRPM3 shows potential as a future therapeutic target for migraine, but further research is required to confirm its role in migraine pathophysiology.