The role of macrophages and iNOS in the effects of CGRP in preclinical migraine models

Editor’s note: The research described below comes from a recipient of a 2024 MSC Travel Grant supporting travel to the 66th Annual Scientific Meeting of the American Headache Society. These grants reimburse travel expenses for those who have had their abstract for a presentation or poster accepted at a meeting.

By Shiva Nematgorgani, PhD student, University of Texas at Dallas, US.

What is the research gap that your study addresses?

Although the role of calcitonin gene-related peptide (CGRP) in migraine pathophysiology has been established, the specific downstream mechanisms and molecular interactions mediated by CGRP are not well understood. Previous studies have highlighted CGRP’s significant impact in triggering migraine through the activation of the trigeminovascular system, leading to vasodilation and neurogenic inflammation. However, the exact molecular pathways through which CGRP influences these migraine-related biological processes, particularly its interactions with immune cells like macrophages and the role of inducible nitric oxide synthase, remain unclear. This gap in understanding hinders the development of targeted therapies that could effectively manage or prevent migraine symptoms.

What is your research hypothesis?

In our model, we hypothesized that the therapeutic potential of alleviating facial hypersensitivity in migraine lies in targeting iNOS and macrophages to mitigate CGRP-induced pain.

What methodology did you use to address your research hypothesis?

Adult CD-1 female mice aged 4-8 weeks were administered intraperitoneal injections of clodronate liposomes (150 μl, IP) to deplete macrophages, or AR-C102222 hydrochloride (15 mg/kg, IP) as a selective iNOS inhibitor followed by dural application of CGRP (1 pg) to induce migraine-like behavioral responses. In the first experiment, mice received injections of clodronate liposomes at 48-hour intervals, followed by dural injections of CGRP or vehicle 24 hours after the second injection of clodronate liposomes or vehicle. In the second experiment, one hour after the administration of AR-C102222 or vehicle, the animals were subjected to dural injections of either CGRP or vehicle. Periorbital mechanical thresholds were assessed using Von Frey filaments at multiple time points post-CGRP.

What are the main results of your study?

The dural application of 1 pg of CGRP resulted in significantly lower facial withdrawal thresholds in female mice. Repeated administration of clodronate liposomes, a macrophage-depleting agent, 24 hours prior to the injection of CGRP, significantly attenuated facial hypersensitivity, suggesting that macrophages may block downstream effects of CGRP.

Additionally, the intraperitoneal injection of AR-C102222 hydrochloride, an iNOS inhibitor, prior to CGRP administration significantly reduced facial hypersensitivity. This suggests that iNOS could play a role in the response to CGRP, inducing facial hypersensitivity.

These findings indicate that iNOS and macrophages may play a critical role in mediating the effects of CGRP in migraine.

What conclusions did you reach based on your results?

This study demonstrated a significant attenuation of dural CGRP-induced behavioral responses by clodronate liposomes or AR-C102222 in female mice. These findings suggest that macrophages and iNOS contribute to the downstream effects of CGRP in migraine.

What are the limitations of your study?

A limitation of this study is the lack of precise quantification of the remaining population of macrophages in the dura following clodronate injection. Another limitation is that the study did not investigate the presence of iNOS-positive macrophages in the dura after exposure to CGRP. All these limitations will be addressed in future experiments.

What is the relevance of your study to migraine?

This study adds to the body of knowledge by elucidating the mechanisms by which CGRP may lead to migraine. This is crucial because understanding the pathways can lead to the development of more targeted migraine treatments that can potentially have greater efficacy than current options. The findings suggest that macrophages and iNOS may be new targets for the development of migraine treatments. This could lead to the development of new drugs or therapeutic strategies to alleviate migraine symptoms in patients.