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Advances in Understanding Migraine: New Mechanisms and Innovative Translational Models

A headache symposium at the 2019 American Pain Society Scientific Meeting highlighted emerging areas of research in the field

by Francie Moehring


16 July 2019


PRF News

MigraineFeatured

A headache symposium at the 2019 American Pain Society Scientific Meeting highlighted emerging areas of research in the field

Migraine is the second most disabling condition worldwide, trailing only low back pain, yet it remains neglected as a field of research. To help bridge the gap, this year’s American Pain Society Scientific Meeting, which took place April 3-6, 2019, in Milwaukee, US, highlighted headache research in a session titled “Studying Headache from Mouse to Human: New Mechanistic Insights and Novel Translational Models.” The take-home message was that while investigators are making progress, more research is needed to fully understand the causes of migraine headache in order to develop better targeted treatments for migraineurs.

 

Looking to hormones to explain sex differences in migraine

The first speaker at the session was Greg Dussor, University of Texas at Dallas, US, who presented his unpublished work on the role of hormones in migraine. “Migraines are two to three times common and more disabling in women than in men,” Dussor said. Interestingly, he noted that the prevalence of migraine in women changes across the menstrual cycle, trimesters of pregnancy, and menopause, suggesting a hormonal component to the condition. However, the exact hormones and mechanisms driving migraine are unknown.

 

Several previous studies suggest that high levels of prolactin, a hormone that signals the body to produce breast milk, could be a contributing factor to migraine (Bosco et al., 2008Cavestro et al., 2006Kallestrup et al., 2014). To test this idea, Dussor and his laboratory first investigated if the prolactin receptor is expressed in nerve fibers that innervate the dura (the outermost meningeal layer surrounding the brain), since dural fibers are thought to be one of the sources of migraine pain. He showed that the prolactin receptor was only expressed in dural nociceptors in female mice and not in male animals. Furthermore, prolactin increased the excitability of female but not male mouse dural afferents. Together, these data suggest that prolactin might be a hormone that contributes to sex differences in migraine.

 

Because increased levels of calcitonin gene-related peptide (CGRP) are associated with migraine, Dussor also measured the amount of CGRP released upon stimulation of isolated rat dura with mustard oil, and assessed how prolactin might contribute to this response. Mustard oil evoked a significantly higher release of CGRP in females than in males pretreated with prolactin, whereas levels of CGRP did not differ significantly between males and females pretreated with vehicle.

 

Prolactin causes facial hypersensitivity in females in a preclinical model of migraine

Next, Dussor introduced a novel preclinical model of migraine that his laboratory has developed (Burgos-Vega et al., 2019). For this minimally invasive model, a cannula is inserted at the intersection of the sagittal and lambdoidal sutures before the skull plates fuse, and migraine-inducing agents are injected directly onto the dura of a mouse.

 

Using this technique, Dussor showed that prolactin elicited facial hypersensitivity (lower facial withdrawal thresholds in response to von Frey filaments applied to the face) and increased grimace scores in female, but not male, mice. Furthermore, the researchers also used the inflammatory agent interleukin-6 (IL-6) to put animals into a “primed” state in which they are more susceptible to future subthreshold doses (doses that in normal controls do not cause pain) of pain-eliciting agents. They found that IL-6 primed female mice to low doses of prolactin, with the animals exhibiting facial hypersensitivity as well as increased grimace scores. Together, the data suggest that small changes in prolactin levels can trigger migraine-like behaviors in female mice.

 

Stress as a trigger

Dussor and his laboratory are always looking for more natural ways to induce migraine-like phenotypes in mice. Because stress is one of the most common migraine triggers, Dussor sought to determine if stress could cause priming to nitric oxide donors (a known migraine trigger in patients). The researchers exposed animals to restraint stress for two hours a day for three days. During this time, the animals exhibit facial hypersensitivity up to 10 days following the last stress-inducing day. Stressed animals that then received sodium nitroprusside (SNP, a nitric oxide donor) exhibited significantly greater facial hypersensitivity than non-stressed control animals. Thus, stress was sufficient to induce in the animals a primed state, which in turn made them more susceptible to a subthreshold trigger later on.

 

When measuring prolactin levels in stressed males and females, Dussor’s group found that stress increased blood levels of prolactin only in females and not in males. To see if prolactin was responsible for the facial hypersensitivity driven by stress, the investigators inhibited prolactin release from the pituitary using bromocriptine. Hypersensitivity levels of stressed animals that received bromocriptine returned to baseline faster and had attenuated priming responses to SNP compared to stressed vehicle-treated controls.

 

Dussor and colleagues also used animals in the stress paradigm in which the prolactin receptor was knocked out from Nav1.8-expressing nociceptors. The researchers found that knockout of the prolactin receptor had no effect in male knockouts, but it abolished the priming response to NO donors after stress in female knockouts compared to wild-type controls.

 

All in all, the findings suggest that prolactin likely contributes to migraine differentially between females and males, and that the prolactin receptor might be a valuable, novel therapeutic target. Furthermore, inhibiting the release of prolactin from the pituitary could potentially be investigated as a female-specific migraine pain treatment.

 

“Yes, we most likely do need female-specific migraine drugs,” Dussor concluded his talk, “and we need to understand the sex-specific actions of drugs before they make it to clinical trials, as choosing the wrong sex may lead to trial failure.”

 

The role of CGRP in migraine

Next in the session was Andrew Russo, University of Iowa, Iowa City, US, who presented his work on CGRP and migraine. CGRP levels are elevated in migraine, and current treatments such as triptans reduce CGRP levels. Furthermore, multiple promising CGRP monoclonal antibodies received US Food and Drug Administration (FDA) approval last year. However, the big remaining question is where CGRP acts in migraine: Is it in the peripheral nervous system (PNS) or in the central nervous system (CNS)?

 

To answer this question, previously Russo and colleagues injected CGRP intraperitoneally (IP) to investigate peripheral mechanisms, or intracerebroventricularly (ICV) to investigate central mechanisms, in wild-type mice (Mason et al., 2017). Then the animals were placed in a two-chamber apparatus where one side was illuminated with bright light and the other with dim light. Russo’s group found that both IP and ICV injections of CGRP caused aversion to bright light, which was attenuated by triptans. This suggested that CGRP could act on both peripheral and central targets.

 

In this same work, Russo and colleagues also turned to a genetic strategy to investigate the PNS versus CNS actions of CGRP. Here, they overexpressed receptor activity-modifying protein 1 (RAMP1), a subunit of the CGRP receptor, in both the CNS and the PNS. As predicted from previous data, in dim light CGRP receptor-overexpressing mice exhibited enhanced light aversion after an ICV (central) injection of CGRP, suggesting that CGRP acts on neurons in the CNS. However, when the researchers performed the same experiment following IP (peripheral) injection of CGRP, they did not observe the same sensitized light aversion. One implication of the findings is that peripheral CGRP may act distinctly from central CGRP, and via a non-neuronal mechanism, to drive the light aversion.

 

Smooth muscle and CGRP

Because peripheral CGRP did not seem to be acting on neurons, Russo next turned to blood vessels and the smooth muscle within them, as CGRP can act directly on blood vessels and lead to vasodilation. In unpublished work, the researchers created mice that overexpressed RAMP1 in blood vessel smooth muscle tissue. These animals exhibited enhanced light aversion compared to littermate controls, even in the absence of exogenous CGRP, and there was a trend to spend even less time in the light upon an IP injection of CGRP (although this was not statistically significant). This enhanced light aversion suggested that peripheral CGRP was most likely acting on vascular smooth muscle cells.

 

To further investigate if the CGRP-induced light aversion was in fact due to vasodilation, as opposed to other actions at the vascular cells, the investigators administered the vasoconstrictors phenylephrine and endothelin-1. Interestingly, blunting the effect of CGRP on systemic blood pressure with phenylephrine had no effect on CGRP-induced light aversion, while endothelin-1 partially attenuated the light aversion. This lack of a clear rescue indicates that peripheral CGRP causes light aversion by mechanisms beyond vasodilation alone.

 

Grimacing and squinting

Russo also sought a more translatable light sensitivity test than the light place aversion assay, so he asked if mice, similarly to humans, would blink in response to a flash of light. However, Russo said, “the animals would sit there with a deer-in-the-headlights kind of look rather than blinking in response to the bright light, [and it] doesn’t matter if the animals were given CGRP or not.”

 

While the animals did not grimace in response to the flash of light, they did show increased grimace scores in response to an IP injection of CGRP compared to saline-injected animals (Rea et al., 2018). Furthermore, the researchers developed a squint assay using video-based measurement of the eyelid fissure—the space between the two open eyelids. Using this squint assay, the researchers showed that CGRP caused squinting in both in darkness and in bright light.

 

In summary, the data show that peripheral CGRP acts on vascular smooth muscle cells, and that peripheral and central CGRP administration lead to light aversion in the animals, similar to what is seen in patients. Furthermore, peripheral CGRP also leads to spontaneous pain that can be measured in the grimace and squint assays, which are both valuable tools for testing novel migraine therapeutics.

 

Migraine photophobia

Concluding the headache session was neurologist Melissa Cortez, University of Utah, Salt Lake City, US, who studies autonomic function and pupillary light responses in migraineurs. Photophobia (sensitivity to light) is a symptom in up to 80 percent of migraineurs during an attack; light can trigger or exacerbate the headache.

 

Previous research from her laboratory has shown that chronic migraineurs exhibit significantly higher light sensitivity as measured by quantitative light sensitivity thresholds, which are associated with altered autonomically-mediated pupillary responses to light, compared to non-headache controls (Cortez et al., 2017). Furthermore, chronic headache (migraine or post-traumatic headache) patients also exhibit systemic autonomic dysfunction, such as gastrointestinal, bladder, and orthostatic dysfunction (Howard et al., 2018), indicating autonomic dysfunction outside the craniofacial domain.

 

Photophobia has been hypothesized to result from a number of potential brain circuit mechanisms, including cortical mechanisms and altered ocular outflow, as well as intrinsically photosensitive retinal ganglion cell pathways acting on dura-sensitive thalamocortical neurons; the latter could affect pupillary responses to light. So Cortez asked if there was a connection between the sensory disruptions of photophobia and craniofacial autonomic dysfunction by examining pupillary function (Cortez et al., 2017).

 

Specifically, Cortez and her laboratory quantitatively assessed photophobia and pupillary responses to light in non-headache controls, probable migraine patients, episodic migraine patients (less than 15 migraines a month), and chronic migraineurs (greater than 15 migraines a month). She found that photophobia thresholds differed among these groups, with chronic migraineurs having a significantly lower photophobia threshold (as measured by the lux, or brightness, of light at which pain is induced) compared to the other groups. Furthermore, Cortez separated groups according to disease severity using clinical measures of headache impact and headache-related disability. Here, Cortez found that photophobia thresholds and pupillary velocities were significantly lower in those with higher headache severity scores.

 

Next, she noted that differences in pupillary diameter changes were associated with the photophobia thresholds across the disease groups examined. In this case, the lowest photophobia thresholds were associated with the largest end pupil size at the photophobia threshold and thus lowest overall pupil constriction change. Of note, too, is that there was a gradient where the association was strongest in chronic migraineurs, second strongest in episodic migraineurs, and weakest in the probable migraineurs. Cortez also saw that those with the most severe migraines had altered pupillary light reflex kinetics, including reduced parasympathetic (pupil constriction latency) and sympathetic pupillary function.

 

Together, the data suggest a possible mechanistic link between sensory sensitization and autonomic homeostasis in migraine.

 

Next, Cortez showed data from work published earlier this year investigating pupillary responses in migraine patients (Cortez at al., 2019). Here, the investigators looked at the pupil cycle time (the time it takes for the pupil to constrict and dilate in response to a slit lamp-based light stimulus); this is a simple pupillary autonomic reflex that has been shown to increase in autonomic neuropathy (Martyn and Ewing, 1986). All the migraine subjects (probable, episodic, and chronic) exhibited longer pupil cycle times, which correlated with a higher number of craniofacial autonomic symptoms, linking pupillary circuit dysfunction to peripheral trigeminal sensitization.

 

Overall, Cortez and colleagues’ work provides the first physiological measure able to distinguish migraine patients from healthy, non-headache controls. Also, the pupillary response to light appears to be an indicator of autonomic circuit dysfunction in migraineurs, which may make it a useful potential biomarker.

 

Francie Moehring, PhD, is a postdoctoral research fellow at the Medical College of Wisconsin, Milwaukee, US.

 

Image credit: Maximilian77/Wikimedia Commons/Creative Commons Attribution-ShareAlike 4.0 International license.

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