**6. New Model of Mechanosensation in Meninges during Migraine Attack**

Based on the concept of the meningeal trigeminovascular system (TGVS) as the initial site for the generation of migraine headache [21], we propose the following model potentially explaining the mechanism of pulsating migraine pain. Figure 2 shows that in interictal state (or in healthy subjects), meningeal nerves express a plethora of pain transducing channels such as mechanosensitive Piezo1 and Piezo2 receptors [5], along with capsaicin-sensitive TRPV1 [36] and ATP-activated P2X3 receptors [37], which all are in low-active non-sensitized state. The main key components of the TGVS, such as meningeal vessels and neighboring nerves, in this state, have a low chance to interact to each other either physically or chemically and a low probability to generate pain signals.

The key event in migraine attack, is the release of the main migraine mediator CGRP [33], which has multiple actions in the TGVS (Figure 2, right). Thus, CGRP induces dilation of meningeal vessels, it promotes local neurogenic inflammation and degranulates dural mast cells, which, in turn, release several pro-inflammatory and pro-nociceptive compounds such as serotonin, histamine, ATP, prostaglandins, and nitric oxide [38–40]. Notably, most of these compounds are able to increase the sensitivity of meningeal afferents to mechanical stimuli [22,41]. Among other compounds released from mast cells, serotonin appeared to be the most strong and fast trigger of nociceptive spiking in nerve terminals [42,43]. Apart from the immediate firing of nociceptors, serotonin also promotes neuronal sensitization and local inflammation directly or through the additional release of CGRP [42]. The inflammation induced by CGRP, substance P and mediators of mast cells can sensitize not only peripheral but also central neurons, expanding the enhanced mechanical sensitivity to extracranial body regions, presented as the phenomenon of allodynia [22,44].

Notably, Piezo1 channels are expressed not only in neurons but also in vessels. Vasodilatation, shear stress and enhanced pulse waves in dilated vessels can activate mechanoreceptors in endothelial cells, triggering, via pannexins, ATP release (Figure 2) [45]. ATP is a strong promoter of nociceptive firing in meninges by itself [37,43] but it is also a trigger of mast cell degranulation promoting further release of the pro-inflammatory and pro-nociceptive compounds to meninges.

**Figure 2.** Schematic presentation of the key elements of the trigeminovascular system comprising meningeal blood vessels, local mast cells and trigeminal nerve fibers before and during migraine attack. **Left:** In the interictal state, before attack, there are only slight pulsations of meningeal vessels with the minimal activation of vascular Piezo1 channels or Piezo1 and Piezo2 channels in nerve fibers. **Right:** During migraine attack, which is often associated with brain oedema and CGRP-induced dilation of vessels, the extracellular space is reduced, promoting more close contact between pulsating vessels, nerves and nearby mast cells. The shear stress in dilated vessels and enhanced vascular pulsations promote mechanosensitive ATP release from the endothelial cells. Mechanical stimulation of calcium permeable Piezo channels in nerve fibers by pulsating vessels promotes neuronal CGRP release. CGRP and ATP can degranulate mast cells directly. In addition, the fraction of mast cells contacting vessels, is directly mechanically activated by blood pulsations. Activation of mast cells induces release of a plethora of pro-nociceptive compounds such as serotonin, histamine, leukotrienes, prostaglandins, ATP, and nitric oxide, further exciting the nociceptive fibers and promoting more CGRP release. All these pro-inflammatory compounds, in long run, together with CGRP, promote neuroinflammation, neuronal sensitization leading to long-lasting pulsating pain.

Approximately one third of migraine cases is represented by migraine with aura. The pathophysiological mechanism underlying migraine aura is a phenomenon called 'cortical spreading depression' (CSD) [46,47]. CSD is a wave of strong depolarization of cortical neurons and glial cells, leading to meningeal neurogenic inflammation, involving the neuropeptide CGRP and substance P, ATP, and the mast cells activation [41,48]. Brain oedema, associated with CSD [46,49] compresses the extracellular space, which assists in the formation of more close contact between blood vessels and meningeal nerve fibers. Furthermore, CSD can increase the brain volume and raise the intracranial pressure [46]. Taken together, these factors, along with strongly pulsating vessels, should facilitate activation of mechanosensitive channels, such as Piezo ones, in nerve fibers [5].

Since meningeal tissues are protected by the skull from the external mechanical forces, the only source for activation of mechanosensitive Piezo channels in meningeal nerves are internal triggers, such as pulsating dural vessels. These pulsating dilated vessels can provide the regular Piezo-mediated excitation of nerve fibers processed towards the high-pain centers and perceived as a pulsating migraine pain. Notably, in migraine condition, there is an increased expansibility of arteries [50], which is also consistent with our hypothesis as the factor supporting the increased amplitude of pulsating waves. The exaggerated mechanical sensitivity of the trigeminal nerves during migraine attack can explain also the painful sensitivity to slight movements of the head or to cough [41].
