Trigeminovascular calcitonin gene-related peptide function in Cacna1a R192Q-mutated knock-in mice

Abstract

Familial hemiplegic migraine type 1 (FHM1) is a rare migraine subtype. Whereas transgenic knock-in mice with the human pathogenic FHM1 R192Q missense mutation in the Cacna1a gene reveal overall neuronal hyperexcitability, the effects on the trigeminovascular system and calcitonin gene-related peptide (CGRP) receptor are largely unknown. This gains relevance as blockade of CGRP and its receptor are therapeutic targets under development. Hence, we set out to test these effects in FHM1 mice. We characterized the trigeminovascular system of wild-type and FHM1 mice through: (i) in vivo capsaicin- and CGRP-induced dural vasodilation in a closed-cranial window; (ii) ex vivo KCl-induced CGRP release from isolated dura mater, trigeminal ganglion and trigeminal nucleus caudalis; and (iii) peripheral vascular function in vitro . In mutant mice, dural vasodilatory responses were significantly decreased compared to controls. The ex vivo release of CGRP was not different in the components of the trigeminovascular system between genotypes; however, sumatriptan diminished the release in the trigeminal ganglion, trigeminal nucleus caudalis and dura mater but only in wild-type mice. Peripheral vascular function was similar between genotypes. These data suggest that the R192Q mutation might be associated with trigeminovascular CGRP receptor desensitization. Novel antimigraine drugs should be able to revert this complex phenomenon.

Keywords: Calcitonin gene-related peptide; R192Q; migraine; trigeminovascular; vasodilation.

Figure 1.

A trace obtained during an experiment on the closed-cranial window. The upper red line represents the dural artery diameter (in arbitrary units, AU) and the lower blue line represents the blood pressure (in mm Hg) simultaneously measured in the femoral artery. The gray vertical lines represent the moment of intravenous administration of endothelin-1 (ET-1, 2 µg/kg), capsaicin (30 µg/kg) or CGRP (10 µg/kg).

Figure 2.

Trigeminovascular dural artery vasodilation. Diameter change (%) induced by endothelin-1 (ET-1), capsaicin or CGRP in wild-type (WT, white bars) and R192Q (grey bars) mice. ET-1 induced comparable dural vasoconstriction in all groups (upper left and right panels). Capsaicin induced significant dural vasodilation in both genotypes (upper left panel), while CGRP only induced dural vasodilation in the wild-type group (upper right panel). The vasodilation induced by capsaicin and CGRP, corrected for the ET-1 baseline, is significantly lower in the R192Q groups than in the wild-type groups (lower left and right panel). Data are expressed as mean ± SEM, n = 9–16, *p < 0.05 vs. the corresponding dose to ET-1, #p < 0.05 between genotypes.

Figure 3.

Effect of pharmacological intervention on mean arterial pressure. Mean arterial pressure (MAP; mm Hg) after administration of endothelin-1 (ET-1), capsaicin or CGRP in wild-type (white bars) and R192Q (grey bars) mice. ET-1 increased the MAP compared to the baseline in both genotypes, and this increase was reverted after administration of only CGRP. Data are expressed as mean ± SEM, n = 9–14, *p < 0.05.

Figure 4.

CGRP release in the trigeminovascular system. Relative stimulated CGRP release in the trigeminal nucleus caudalis (TNC), trigeminal ganglion (TG) and dura mater of wild-type (WT, white bars and circles) and R192Q (grey bars and squares) mice. There is no statistical difference in CGRP release between wild-type and R192Q mice in the trigeminal nucleus caudalis, trigeminal ganglion or dura mater. Data are expressed as mean ± SEM, n = 11–12.

Figure 5.

Effect of repeated stimulation and sumatriptan on CGRP release. Relative stimulated CGRP release after a first (1^st) and second (2^nd) stimulation with KCl (panels a, b, c) or in the absence (−) or presence (+) of sumatriptan (panels d, e, f) in trigeminal nucleus caudalis (TNC), trigeminal ganglion (TG) and dura mater of wild-type (WT, circles) and R192Q (squares) mice. There were no statistically significant differences between the first and second stimulation with KCl in either genotype. Sumatriptan significantly reduced the relative stimulated CGRP release in the trigeminal nucleus caudalis, dura mater and trigeminal ganglion of wild-type, but not in all the trigeminovascular components of the R192Q mice tested. Data are expressed as mean ± SEM, n = 5–6, *p < 0.05 vs. response in the absence of sumatriptan.

Figure 6.

Peripheral vascular function. Concentration-response curve to different agonists in isolated aortas and mesenteric arteries of wild-type (WT, open circles) and R192Q (closed circles) mice. There were no differences between wild-type and R192Q mice in relaxant responses to acetylcholine (a, e), sodium nitroprusside (SNP, b, f) and CGRP (c, g), which are expressed relative to the contraction induced by U46619, in the aortas (a, b, c) and mesenteric arteries (e, f, g). Contractile responses to 5-hydroxytryptamine (5-HT, d, h), which are expressed relative to 100 mM KCl, in the aortas (d) and mesenteric arteries (h) were also not different between wild-type and R192Q mice. Data are expressed as mean ± SEM, n = 10–13.