*Review* **Migraine: Advances in the Pathogenesis and Treatment**

**Horia Ples, 1 , Ioan-Alexandru Florian 2,\* , Teodora-Larisa Timis 3,\* , Razvan-Adrian Covache-Busuioc <sup>4</sup> , Luca-Andrei Glavan <sup>4</sup> , David-Ioan Dumitrascu <sup>4</sup> , Andrei Adrian Popa <sup>4</sup> , Andrei Bordeianu <sup>4</sup> and Alexandru Vlad Ciurea <sup>4</sup>**


**Abstract:** This article presents a comprehensive review on migraine, a prevalent neurological disorder characterized by chronic headaches, by focusing on their pathogenesis and treatment advances. By examining molecular markers and leveraging imaging techniques, the research identifies key mechanisms and triggers in migraine pathology, thereby improving our understanding of its pathophysiology. Special emphasis is given to the role of calcitonin gene-related peptide (CGRP) in migraine development. CGRP not only contributes to symptoms but also represents a promising therapeutic target, with inhibitors showing effectiveness in migraine management. The article further explores traditional medical treatments, scrutinizing the mechanisms, benefits, and limitations of commonly prescribed medications. This provides a segue into an analysis of emerging therapeutic strategies and their potential to enhance migraine management. Finally, the paper delves into neuromodulation as an innovative treatment modality. Clinical studies indicating its effectiveness in migraine management are reviewed, and the advantages and limitations of this technique are discussed. In summary, the article aims to enhance the understanding of migraine pathogenesis and present novel therapeutic possibilities that could revolutionize patient care.

**Keywords:** migraine pathogenesis; molecular markers; calcitonin gene-related peptide (CGRP); migraine treatment; neuromodulation

### **1. Introduction**

An overwhelming majority of the global population, approximately 95%, have suffered from a headache at some point in their lives, with an alarming annual prevalence that suggests nearly half of all adults have experienced a headache within a given year [1]. The ramifications of this health issue extend far beyond personal discomfort, with headaches accounting for one-tenth of consultations with general practitioners [2], and a significant portion, one-third, of referrals to neurologists [3]. Moreover, acute medical admissions related to headaches are alarmingly high, constituting one in every five cases.

The World Health Organization recognizes the debilitating nature of headaches, including them among the top ten global causes of disability. Interestingly, in women, the prevalence and impact of headaches are even more pronounced, ranking among the top five causes of disability [4]. It is pertinent to note that the debilitating impact of headaches

**Citation:** Ples, , H.; Florian, I.-A.; Timis, T.-L.; Covache-Busuioc, R.-A.; Glavan, L.-A.; Dumitrascu, D.-I.; Popa, A.A.; Bordeianu, A.; Ciurea, A.V. Migraine: Advances in the Pathogenesis and Treatment. *Neurol. Int.* **2023**, *15*, 1052–1105. https:// doi.org/10.3390/neurolint15030067

Academic Editor: Yasushi Shibata

Received: 1 August 2023 Revised: 28 August 2023 Accepted: 28 August 2023 Published: 31 August 2023

**Copyright:** © 2023 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

is comparable to chronic conditions, such as arthritis and diabetes, and its severity exceeds that of conditions like asthma [5,6].

Taking the United Kingdom as an illustrative example, the socioeconomic implications of migraine, a specific type of headache, are vast. An estimated 25 million workdays are lost annually due to migraine, creating an indirect economic burden of nearly GBP 2 billion per year. This figure does not even include the direct healthcare costs associated with managing headaches, such as medication expenses, consultations with general practitioners, referrals to specialists, and visits to emergency care facilities [7].

Quantifying the influence of headaches, particularly migraine, on an individual's quality of life can be challenging. Yet, it is clear from reported data that the impact is substantial. A significant percentage, approximately 75% of patients, experience functional disability during a migraine attack. Additionally, half of the sufferers require the assistance of family members or friends during an attack, causing a significant disruption to their social lives [8]. The ripple effect of headaches extends beyond the individuals suffering from them, impacting society at large and requiring serious attention for more effective management strategies [9].

Migraine, a long-term headache disorder punctuated by episodic bouts, is characterized by repeated instances of severe headaches that present with unique associated symptoms. These include photophobia, a heightened sensitivity to light, and phonophobia, an increased sensitivity to sound [10]. The classification of episodic migraine—an intermittent but recurring form of this disorder—hinges on the frequency with which a patient experiences these debilitating headaches.

In the majority of cases, patients undergo fewer than 15 episodes of headaches per month, a condition identified as episodic migraine. Conversely, there is a subset of individuals who face a more frequent occurrence of headaches—on 15 or more days each month, spanning over three months. Importantly, at least eight of these days should either meet the diagnostic criteria for migraine without the accompanying aura or show responsiveness to treatment specifically designed for migraine. The International Headache Society recognizes this latter classification as chronic migraine [11].

Chronic migraine, although less common when compared to its episodic counterpart, remains a pervasive and incapacitating issue [12]. It poses a significant burden on those afflicted with the condition, dramatically impacting their daily lives and well-being [13,14]. This persistent form of migraine continues to be a widespread challenge, necessitating ongoing research and improved therapeutic strategies to ease the strain it puts on sufferers [15].

#### *1.1. Brief Overview of Migraine as a Prevalent Neurological Condition*

Over the past three decades, there has been a significant upsurge in the worldwide prevalence of migraine. As highlighted by the Global Burden of Disease (GBD) 2019 study, the estimated global occurrence of migraine escalated from 721.9 million (with a 95% uncertainty interval (UI) of 624.9–833.4) in 1990 to a staggering 1.1 billion (95% UI: 0.98–1.3) in 2019. The percentual shifts in global age-standardized prevalence rate and years lived with disability (YLDs) over these nearly three decades were recorded at 1.7 (95% UI: 0.7–2.8) and 1.5 (95% UI: −4.4 to 3.3), respectively [16].

In this time frame, the sharpest escalations in the age-standardized prevalence per 100,000 individuals were recorded in East Asia with a 7.9% increase (95% UI: 4.3–12%), and in Andean Latin America with an increase of 6.7% (95% UI: 2.1– 11.9%). Conversely, the most significant decreases were seen in high-income North America [−2.2% (95% UI: −5.3 to 1.1%)] and Southeast Asia [−2.2% (95% UI: −3 to −1.4%)]. Moreover, the age-standardized YLD rate due to migraine also saw an increase from 517.6 (95% UI: 82.0–1169.1) in 1990 to 525.5 (95% UI: 78.8–1194.0) in 2019.

The incidence of migraine consistently appeared higher in females than in males across all age groups. In 2019, the global age-standardized prevalence rate for females was 17,902.5 (95% UI: 15,588.3, 20,531.7) per 100,000 populations, in comparison to 10,337.6 (95% UI: 8948.0, 12,013.0) for males [16].

Notably, the most frequent incidence of migraine, both in terms of rate and absolute number of new cases, was seen in the age bracket of 10–14 years for both genders. Over the course of 2019, the number of YLDs due to migraine began to increase from birth, reaching a peak in the 30–34 age group, after which it slowly receded for both sexes [17].

Contrary to expectations, socioeconomic status did not appear to have a direct correlation with the burden of migraine. The study did not reveal any discernible link between the socio-demographic index (SDI) and the YLD rate associated with migraine. This lack of association suggests that migraine does not discriminate based on socioeconomic status, further underlining the pervasive nature of this debilitating condition [17].

The prevalence of migraine has been reported to fluctuate between 2.6% and 21.7%, with an estimated average prevalence close to 12%. However, these figures vary significantly across different nations and even between individual studies conducted within the same country [18–21].

Notably, there appears to be a strong familial connection among individuals suffering from migraine, suggesting that genetic factors significantly contribute to the risk of developing this condition [20,22–25]. Supporting this theory, twin studies have indicated that migraine represent a complex genetic disease that involves an intricate interplay between genetic and environmental factors. Remarkably, the heritability of migraine has been estimated to be as high as 65% [26–29].

However, in spite of the robust genetic implications suggested by these studies and several large-scale genome-wide association studies (GWAS) conducted over the years, the scientific community has yet to conclusively identify specific candidate genes responsible for migraine. A recent systematic re-evaluation of 27 proposed candidate genes found none to be statistically significant [30].

Interestingly, the prevalence of migraine among neurologists is significantly higher when compared to the general population, reaching prevalence rates as high as 48.6% in some studies. This elevated prevalence is most likely attributable to enhanced selfrecognition of migraine symptoms among professionals who are extensively trained and experienced in diagnosing and treating the condition. This assertion is supported by a study revealing that just over half of individuals who were diagnosed with migraine actually recognized their headache as a migraine [31].

#### *1.2. Prevalence of Migraine in Pediatric Patients*

From a meta-analysis, in which data were sourced from 40 studies encompassing a sample of 15,626 pediatric and adolescent individuals diagnosed with migraines, an 11% prevalence rate was noted, displaying considerable heterogeneity. Among these, 27 studies delineated migraine prevalence based on gender. The aggregated prevalence rate for females stood at 4%, whereas for males it was 3%. Specific data concerning MwoA (migraine without aura) and MwA (migraine with aura) were gleaned from 13 studies, which covered 3481 and 1322 subjects diagnosed with MwoA and MwA, respectively. Prevalence for MwoA was identified at 8% and for MwA at 3%, with marked heterogeneity for both. Only six studies offered data on chronic migraines, revealing a prevalence that fluctuated between 0.2% and 12% [32].

From a separate dataset of 31 studies, information was extracted involving 13,105 pediatric and adolescent subjects diagnosed with TTH (tension-type headache). This cohort exhibited a prevalence of 17%, with notable heterogeneity. Out of these studies, 23 offered a gender-based breakdown of TTH prevalence, yielding a consolidated prevalence rate of 11% for females and 9% for males. Limited data on episodic and chronic TTH were derived from 7 studies, which presented a prevalence range of 4–29% and 0.2–12.9%, respectively [32].

Another set of data, obtained from 40 studies, encompassed 76,782 pediatric and adolescent participants diagnosed with primary headaches in general. The overall prevalence was determined at 62%, with significant heterogeneity observed. Gender-based prevalence data for primary headaches, extracted from 29 studies, showed an aggregated prevalence rate of 38% for females and 27% for males [32].

#### *1.3. Medical Treatments of Migraine in Children*

Recent advancements in the pharmaceutical sector have introduced a selective 5-HT1F agonist, lasmiditan, which serves as an efficacious acute treatment for adults, demonstrating no vasoconstrictor activity. This drug is currently under investigation for its applicability in pediatric populations. Additionally, several novel calcitonin gene-related peptide (CGRP) antibodies and antagonists, which have demonstrated efficacy in both the acute treatment and prevention of migraines in adults, are now being assessed in pediatric clinical trials. In adult medical practices, there is an increasing inclination towards peripheral nerve blocks and botulinum toxin; however, the need for robust evidence supporting their efficacy in children is paramount. Furthermore, the introduction of electroceuticals—therapeutic electric devices—has broadened the treatment horizon. These devices include the external trigeminal nerve stimulator (e-TNS), non-invasive vagal nerve stimulator (nVNS), singlepulse transcranial magnetic stimulator (sTMS), and remote electrical neuromodulation device (REN). Presently, substantial evidence supporting their effectiveness in pediatric populations remains elusive; furthermore, while significant progress has been observed, it predominantly benefits the adult demographic. There is an imperative need to expedite migraine research focusing on children [33].

### **2. Pathogenesis of Migraine: Role of Molecular Markers in Identifying Migraine Triggers and Mechanisms**

#### *2.1. Definition and Significance of Molecular Markers*

Biomarkers, in the realm of medical and biological research, are defined as quantifiable indicators of biological conditions, representing either physical manifestations or results obtained from laboratory tests that correlate with biological processes. These markers have the potential to serve critical diagnostic or prognostic functions [34]. A more explicit definition of biomarkers was proposed during a conference hosted by the US Food and Drug Administration. In this context, biomarkers are characterized as quantifiable attributes that can be objectively measured and assessed, providing insights into standard biological, pathological, or pharmacological processes [35].

This clear and precise definition paves the way for a bifurcation of biomarkers into the following two unique types: diagnostic and therapeutic. Diagnostic biomarkers serve as flags for pathological conditions and bear a close association with the risk of developing a disease and its severity. They aid in identifying the presence of a disease and gauging its stage or intensity, thus playing a crucial role in guiding clinical decision-making [36].

On the other hand, therapeutic biomarkers hold a different but equally important role. They provide information on a treatment's response, effectively serving as indicators of the efficacy or success of a therapeutic intervention. These biomarkers help clinicians tailor treatments to individual patients, allowing for personalized medicine approaches. They offer a chance to predict whether a patient is likely to respond positively to a particular treatment, making them a powerful tool in the management and treatment of diseases. By providing an early indication of the effectiveness of a therapeutic regimen, these markers can guide healthcare professionals in adjusting treatments as necessary, minimizing the trial-and-error aspect of disease management and increasing the probability of successful outcomes [15].

Biomarkers represent objective physical traits that can be harnessed to illuminate and distinguish the biological nature and mechanisms of various diseases and syndromes. Essentially, they provide snapshots of the body's physiological state and can offer valuable insights into health and disease processes. Biomarkers have an extensive range of potential manifestations, which can include but are certainly not limited to, results obtained from the examination of blood, urine, muscle, nerve, skin, or cerebrospinal fluid [37].

Additionally, biomarkers may also be identified in the form of genes or gene products. These genetic markers offer a unique insight into an individual's inherent disease susceptibility or resistance and can often illuminate potential therapeutic pathways. Likewise, biomarkers can be identified through advanced imaging techniques such as X-rays, magnetic resonance imaging (MRI), or computed tomographic (CT) scans. These imaging biomarkers can provide a visual representation of disease progression, allowing clinicians to identify anatomical or functional changes in the body over time [34].

Another fascinating domain of biomarkers lies in the realm of electrophysiological measurements, such as those generated by electrocardiograms (ECGs), electroencephalograms (EEGs), or nerve conduction studies. These types of biomarkers record the electrical activity of the heart, brain, or nerves, respectively, offering a unique insight into the physiological function of these systems.

An important issue worth mentioning is those paraclinical investigations offer a new avenue for the management of migraine but are not proven to be of high sensibility and sensitivity for daily physician's practice. Even though neuroimaging and functional analyses of the brain activity might give a broader point of view regarding therapeutic possibilities, those should not be taken into consideration as absolute clinical criteria.

Ultimately, a biomarker could be virtually any characteristic that can be detected, quantified, and expressed in terms of physical qualities. These could include diverse measures, such as height, weight, depth, voltage, luminescence, resistance, viscosity, width, length, volume, or area. Each of these measures contributes to the vast array of biomarkers that hold promise for enhancing our understanding of diseases and guiding the development of effective therapeutic interventions. The utilization of such a wide array of biomarkers allows for a comprehensive, multi-faceted approach to understanding and treating diseases, ultimately leading to more effective and personalized healthcare solutions [38].

#### *2.2. Identification of Potential Molecular Markers Associated with Migraine*

The National Institutes of Health Biomarkers Definitions Working Group, in 1998, presented a definition for biomarkers. As per their definition, a biomarker refers to "a characteristic that can be objectively measured and evaluated as an indicator of normal biological processes, pathogenic processes, or pharmacological responses to a therapeutic intervention" [35]. Biomarkers may be classified based on their functional roles, such as diagnostic, therapeutic, risk, progression, and prognostic indicators.

The 'ideal' biomarker is characterized by the following features [39]:


In the context of migraine, however, there are no validated biomarkers due to the absence of substance or genetic variants that are exclusively associated with this condition or the lack of comprehensive studies on potential biomarkers.

#### 2.2.1. Markers of Inflammation and Oxidative Stress

The markers of inflammation and oxidative stress have been associated with migraine in several studies. Proinflammatory cytokines, such as interleukin-1 (IL-1) and interleukin-6 (IL-6), have been implicated in this condition [40]. It has been found that the level of IL-1α is elevated in the blood of children suffering from migraine with aura (MA) [40]. Similarly, adults with MA have been found to exhibit higher plasma levels of IL-1β during headache-free periods and early stages of attacks as compared to those suffering from migraine without aura (MO) [40,41].

The concentration of IL-6 is reported to increase during the initial two hours of a migraine attack. Additionally, the levels of IL-10 and tumor necrosis factor alpha (TNFα) are also found to be elevated during these attacks. It is believed that other inflammatory markers associated with vascular dysfunction, such as homocysteine (Hcy) and matrix metalloproteinase-9 (MMP-9), are also elevated in the blood of individuals with migraine [15].

Elevated serum Hcy concentration has been linked to migraine with aura (MA), and some studies have noted a relationship between increased Hcy levels and higher frequency and severity of migraine; however, these findings are not supported by all research. Hyperhomocysteinemia (elevated Hcy) is hypothesized to initiate migraine with aura attacks through changes in pain threshold [42,43].

#### 2.2.2. Markers Associated with Pain Transmission and Emotions

Biochemical research has revealed several metabolic irregularities in the synthesis of neuromodulators and neurotransmitters associated with migraine, particularly migraine without aura (MO). Alterations in the metabolic pathway of tyrosine, for example, lead to abnormal production of neurotransmitters like noradrenaline (NE) and dopamine (DA). This process results in an increase in the levels of trace amines, such as tyramine, octopamine, and synephrine. Such changes compromise mitochondrial function and elevate glutamate concentrations within the central nervous system (CNS), as can be seen in Table 1 [43].

These imbalances in the neurotransmitter and neuromodulator levels within the dopaminergic and noradrenergic synapses of pain pathways could potentially activate the trigeminovascular system (TGVS), causing the release of the calcitonin gene-related peptide (CGRP). This chain of events is believed to directly trigger migraine attacks [44,45].

CGRP plays a key role in transmitting pain signals and promoting inflammation. Its release is stimulated by the activation of TGVS and severe migraine episodes. Infusion of CGRP has been observed to provoke migraine-like attacks in patients with migraine with aura (MA). It has been reported that during inter-attack periods, the saliva and plasma levels of CGRP in migraine patients are significantly higher compared to healthy individuals [43].

Research conducted on cultured trigeminal neurons suggests that migraine treatment strategies can inhibit CGRP transcription and curtail its release, while tumor necrosis factor alpha (TNF-α) may stimulate the transcription of this peptide [15]. Another study proposes that high levels of CGRP in saliva may correlate with a significantly improved response to rizatriptan treatment, suggesting that CGRP could serve as a valuable therapeutic marker [46].

Glutamate, which could potentially activate pathways involving both TGVS and cortical spreading depression (CSD), has been found in elevated concentrations in the plasma, platelets, and cerebrospinal fluid (CSF) of migraine sufferers, including those with chronic migraine. Research suggests that a reduction in plasma glutamate levels could be a marker of a positive response to prophylactic treatment in MO patients [43].

Serotonin (5-HT) release from platelets into the plasma may be implicated in the pathophysiology of the aura phase of migraine. Izzati-Zade observed a depletion of 5-HT stored in platelets during migraine attacks; moreover, a pattern has been observed in which the plasma level of 5-HT decreases between migraine attacks and the level of the corresponding metabolite, hydroxyindoleacetic acid (5-HIAA), increases. This pattern reverses during migraine attacks [47,48]. This correlation suggests that low 5-HT levels might enable the activation of the trigeminovascular nociceptive pathway triggered by CSD, thus supporting the hypothesis that migraines are a syndrome of low serotonergic disposition.

Additionally, a significantly higher concentration of hypocretin-1, a wakefulnesspromoting neuropeptide, has been detected in the CSF of patients with chronic migraine, and this has been observed to correlate with painkiller usage [49,50]. Elevated hypocretin-1

levels may be indicative of the early stages of a migraine attack. Conversely, a study involving patients with cluster headaches reported reduced hypocretin-1 levels in the CSF, suggesting that low hypocretin-1 concentrations might reflect insufficient antinociceptive activity in the hypothalamus [51].

New therapeutic targets for migraine treatment, such as CGRP receptor antagonists, anti-CGRP antibodies, 5-HT1F agonists, glutamate antagonists, and dual hypocretin-1 receptor antagonists, are currently under investigation in phase II clinical trials [52,53]. These emerging therapies reflect the continuous exploration and evolution of our understanding of migraine pathophysiology.

**Molecule Migraine Type (Chronic Migraine [CM]/Episodic Migraine [EM]) Action in Relation to Migraine Sodium** [54,55] EM • During a migraine, there is an increase in cerebrospinal fluid (CSF) sodium concentration, while the blood plasma sodium concentration remains unchanged. Additionally, sodium excursions may follow a temporal pattern that worsens migraine in susceptible patients **Homocysteine** [56] EM • High levels of homocysteine are potentially linked to migraine with aura and an increased risk of cardiovascular events in patients with migraine **3,4-Dihydroxyphenylacetic acid (DOPAC)** [57] EM • Related with dopaminergic activity • Positive correlation between the concentration of DOPAC (3,4-dihydroxyphenylacetic acid) and the intensity of migraine, whether with or without aura **Phosphatidylcholine-specific phospholipase C** [58] EM • The process involves the hydrolysis of phosphatidylcholine, resulting in the production of important second messengers, diacylglycerol, and phosphorylcholine **Transforming growth factor-**β**1** [59] EM, CM • An anti-inflammatory cytokine **Interleukin-1 receptor antagonist** [59] EM, CM • Proinflammatory cytokine **Monocyte chemoattractant protein-1** [59] EM, CM • Proinflammatory cytokine **Corticotrophin-releasing factor** [60] CM, MOH • May be involved in activation of hypocretin/orexin system. **Orexin-A (also referred to as hypocretin-1)** [60] CM, MOH • Involved in the maintenance and regulation of various physiological functions, including arousal, sleep, appetite, drinking behavior, central control of autonomic activity, certain endocrine responses, and pain modulation **Glial cell line-derived neurotrophic factor** [61] CM • It may play a role in pain relief by regulating the expression of sodium channel subunits, capsaicin VR1 receptors, and substance P release • Reduced levels found in patients with migraine **Somatostatin** [61] CM • Regulatory anti-inflammatory and antinociceptive peptide **Glutamate** [62] CM • The primary excitatory neurotransmitter in the central nervous system. It has been linked to various migraine-related processes, including cortical spreading depression, trigeminovascular activation, and central sensitization.

**Table 1.** Molecules with altered CSF (cerebrospinal fluid) concentrations in patients with migraine.


#### **3. Other Biomarkers Associated with Increased Risk for Migraine**

*3.1. Genetic Markers and Migraine*

Many scientific investigations have striven to identify specific genetic mutations or polymorphisms that might contribute to an increased risk of developing migraine. However, as of now, none of these findings have been implemented in standard clinical practice. One rare subtype of migraine, known as familial hemiplegic migraine (FHM), which is characterized by aura and transient hemiplegia, has a well-understood genetic basis. There are three known genes where mutations have been linked with FHM—CACNA1A (FHM1), ATP1A2 (FHM2), and SCN1A (FHM3)—and this condition is inherited in an autosomal-dominant fashion [66].

The identified mutations connected to FHM lead to alterations in calcium and sodium channel functions, which are integral components of neuronal communication and excitability. Interestingly, these genetic variants have also been associated with other neurological disorders, including ataxia and childhood epilepsy [66]. Nevertheless, these mutations have not shown a strong correlation with common forms of migraine (with or without aura) or other types of headaches.

A recent study discerned a significant genetic correlation linking migraine risk to intracranial volume (rG = <sup>−</sup>0.11, P = 1 <sup>×</sup> <sup>10</sup>−<sup>3</sup> ). This correlation was not observed in relation to any subcortical region. Notwithstanding, the study pinpointed concurrent genomic overlap between migraines and all brain structures. Gene enrichment in these mutual genomic regions indicated potential associations with neuronal signaling and vascular regulation. Furthermore, the research suggested a potential causative link between reduced overall brain volume, as well as the volume of the hippocampus and ventral diencephalon, and heightened migraine risk. Additionally, a causative correlation was proposed between heightened migraine risk and an expanded amygdala volume. Through the utilization of comprehensive genome-wide association studies, the study illuminated shared genetic pathways influencing both migraine risk and various brain structures. This suggests that variances in brain morphology in individuals with elevated migraine susceptibility could be rooted in genetics. Delving deeper into these findings offers support to the neurovascular premise of migraine origin, highlighting prospective therapeutic avenues [67].

Another study elucidated the following genes associated with familial hemiplegic migraine [68]:



Two FHM1 knock-in (KI) transgenic mouse models have been established as per references [69,70]. The KI model for the R192Q mutation, linked with pure FHM1, does not exhibit clinical anomalies. In contrast, the KI for the S218L mutation, attributed to severe FHM1, presents cerebellar ataxia, transient hemiparesis, and epilepsy. As outlined in [71], these FHM1-KI mice demonstrate heightened CaV2.1 currents and neurotransmitter release, an imbalance in cortical neurotransmission, amplified excitatory transmission in the visual cortex, and a higher vulnerability to cortical spreading depression (CSD).

Various models of FHM2-KI transgenic mice have been developed. Heterozygous transgenic mice [72] display no clinical changes but have an elevated predisposition to CSD. Mice with a partial knock-out (KO) of ATP1A2 also demonstrate a heightened vulnerability to CSD [73]. Another model with a complete KO of ATP1A2 in astrocytes manifests episodic paralysis and spontaneous CSD waves coupled with diminished EEG activity. Aberrations in brain metabolism were observed with increased levels of serine and glycine. Interestingly, a diet devoid of serine and glycine curtailed paralysis episodes in these mutants [74].

For FHM3, multiple SCN1A mutations have been documented, with the majority being missense alterations leading to enhanced function [75]. A mouse model harboring the L1649Q variant exhibited an increased susceptibility to CSD, attributed to Na+ channel inactivation defects and augmented Na+ currents, causing hyperactivity in inhibitory interneurons.

With respect to FHM4, mutations in PRRT2 have been discovered in numerous instances as referenced in [76]. A significant portion of these cases were pure FHM, while others exhibited accompanying epilepsy, cognitive impairments, or dyskinesia. PRRT2-KO mice displayed paroxysmal abnormal movements early in life, progressing to unusual audiogenic motor behaviors in adulthood and a reduced seizure threshold. Notably, both human and mouse homozygous KO-PRRT2 neurons in culture exhibited hyperactive NaV1.2 and NaV1.6 channels, inferring PRRT2's inhibitory effect on voltage-gated sodium channels as described in [77].

Researchers have also employed genome-wide association studies (GWAS) to pinpoint genes linked to an elevated susceptibility for migraine (see Table 2). In one such investigation, genetic information from 5122 individuals afflicted with migraine and 18,108 control participants was scrutinized. This scrutiny led to the identification of several specific genetic variations known as single-nucleotide polymorphisms (SNPs), which displayed significant connections to migraine. Noteworthy among these were rs2651899 (positioned on chromosome 1p36.32, close to the PRDM16 gene), rs10166942 (situated on 2q37.1, near TRPM8), and rs11172113 (positioned on 12q13.3, near LRP1). It is important to highlight that although rs2651899 and rs10166942 could be differentiated between migraine and non-migraine headaches, these three SNPs did not exhibit exclusivity for migraine with or without aura, nor were they tied to specific migraine characteristics. Nonetheless, the biological significance of these connections is substantiated by the established functions of TRPM8 in neuropathic pain and LRP1 in glutamatergic synaptic transmission [78].

Another GWAS pinpointed the following two susceptibility loci for migraine without aura: MEF2D and TGFBR2 [79]. It is important to bear in mind that the results from GWAS carried out have not overlapped so far, and larger-scale studies are necessary to confirm and expand the findings of smaller investigations and to permit the use of meta-analytical methodologies.

A migraine GWAS study from 2021 [80] identified 79 independent loci significantly correlated with migraine. This study was ethnically diverse, encompassing participants of East Asian, African American, and Hispanic/Latino origin, and consisted of 28,852 cases versus 525,717 controls.

The latest migraine GWAS from 2022 by Hautakangas et al. comprised 102,084 cases against 771,257 controls. This study unearthed 123 unique loci associated with migraines, 86 of which were newly discovered post the 2016 GWAS. Further studies even expanded independent SNPs to 167. The 2022 GWAS [81] underscored both vascular and CNS tissues/cell types. Newly detected loci encoded migraine drug targets, such as CGRP (CALCA/CALCB) and serotonin 1F receptor (HTR1F). Significantly, CGRP is the objective for CGRP antibodies, and HTR1F is targeted by ditans. Moreover, an in-depth assessment of roughly 30,000 patients from the 2022 GWAS with a precise migraine diagnosis revealed unique risk variants for specific migraine types.

The research presented thus far suggests that, aside from FHM, we are only at the preliminary stage of identifying genes significantly associated with migraine risk [82]. This observation is further illustrated by the inconsistent findings from studies investigating specific associations in migraine patients with and without aura (summarized in Table 2). For instance, one study found a significant association between a polymorphism in the gene encoding the dopamine D2 receptor (see Table 2) and migraine without aura [83]. Meanwhile, another study supported the association of DBH and SLC6A3 genes with migraine with aura [62]. Contradictorily, other investigations did not corroborate these associations in migraine patients, whether with or without aura. This variability is not unusual in genetic studies investigating diseases with a multifactorial etiology. As such, further research is needed to unravel the complex genetic underpinnings of migraine [15].


**Table 2.** Genetic mutations/polymorphisms associated with increased risk for migraine and relation to migraine.


**Table 2.** *Cont.*

#### *3.2. Recent Genetic Findings and Migraine*

In a newly published family-based association study, significant markers connected to migraine were discovered, alongside genes believed to contribute to or modify the phenotypic expression of migraine within a substantial region of chromosome 6p12.2–p21.1. This region is recognized by the locus name MIGR3. Regrettably, due to the vastness of this area of interest, it is currently not feasible to pinpoint a singular gene; however, it is anticipated that future investigations employing more refined sequencing methodologies will eventually lead to the identification of a promising candidate gene implicated in migraine [98,99].

Despite the growing body of evidence suggesting that genetic factors play a pivotal role in the development of migraine, efforts to uncover the specific genes responsible for the common forms of migraine have only yielded modest success. As scientific collaboration expands on a global scale, the chances of identifying additional genetic variants linked to migraine are likely to increase. Furthermore, the unraveling of the genetic intricacies underlying polygenic diseases could potentially shed new light on the molecular pathways implicated in the pathophysiology of migraine. By extending our understanding of the genetic aspects of migraine, we may pave the way for the development of more effective diagnostic tools and therapeutic interventions.

#### *3.3. Inflammatory Indicators and Migraine*

Interleukins, specifically IL-1 and IL-6, have been linked with the occurrence of migraine. These cytokines, characterized by their proinflammatory nature, are believed to play a role in vascular dysfunction. Studies indicate that children experiencing migraine have raised plasma levels of IL-1α compared to those who do not suffer from migraine, and these concentrations are markedly higher in individuals with migraine accompanied by aura compared to those without aura [40]. Furthermore, adults experiencing aura migraine have significantly elevated plasma levels of IL-1β during periods free from headaches and during the early onset of migraine attacks, in comparison to individuals with migraine that do not present with aura [41]. IL-6 levels also exhibit a surge in the initial two hours of a migraine attack when measured from blood samples taken from the jugular vein [50].

Other cytokines, including IL-10 and tumor necrosis factor alpha (TNF-α), have shown associations with migraine. During migraine attacks, there are elevated serum levels of IL-10 and TNF-α [100]; moreover, between attacks, TNF-α levels in plasma are higher in children who suffer from migraine compared to those who do not. The connection between TNF-α and migraine is particularly noteworthy, given the repeated association of elevated levels of this cytokine with endothelial dysfunction [63]. While some studies propose that patients with migraine may have compromised endothelial function, others contradict these findings [15].

Further inflammatory markers, which are considered to be linked to vascular dysfunction, are found to be elevated in the blood of migraine patients. Research has demonstrated

that average plasma levels of C-reactive protein and homocysteine are higher in children who suffer from migraine compared to those who are not plagued by headaches [54]. Evidence also suggests that premenopausal women with migraine, especially those with aura, show signs of increased endothelial activation—a component of endothelial dysfunction evidenced by elevated levels of von Willebrand factor, C-reactive protein, nitrate/nitrite, and tissue-type plasminogen activator antigen [55]. Markers linked to vascular repair and remodeling processes have also shown an association with migraine.

Investigations in both human subjects and animal models have proposed that matrix metalloproteinase-9 (MMP-9) might protect against the development and destabilization of plaques [61]. Moreover, patients experiencing migraine have been found to have significantly higher plasma levels of MMP-9 compared to healthy individuals and those with tension-type headaches. The average plasma MMP-9 levels were highest in subjects who had their blood samples taken between two and four days post their latest attack, implying that the elevated MMP-9 might be an indication of structural damage and subsequent remodeling associated with migraine attacks [64].

While the findings summarized here point to a relationship between various inflammatory mediators and migraine, additional research aimed at understanding the biological implications of these inflammatory mediators is necessary to confirm the validity of these potential indicators.
