16th European Headache Congress

Prof.

Chair

Effects of amylin on eating and sites of action in the brain

Invited speaker

Invited lecture

Amylin in migraine and pain-related behaviours in animal models

Amylin and migraine in humans

Amylin 1 receptors produce delayed activation and sensitization of trigeminovascular neurons

Abstract text (incl. figure legends and references) Objective: Investigate in an in vivo model the role of amylin 1 (AMY1) receptor activation in the modulation of the trigeminal nociceptive system in rats during the different phases of the estrous cycle, and compare it to the responses observed in males. Methods: We recorded neuronal activity in male and female rats with extracellular electrodes placed within the trigeminocervical complex and examined the effects of targeting the AMY1 receptor on ongoing spontaneous and dural-evoked firing rates of central trigeminovascular neurons. The selective AMY1 receptor agonist pramlintide and AMY1 receptor antagonist AC187, were used for the present study. The different stages of the estrous cycle were identified and assigned by a blinded experimenter through Cresyl violet-stained vaginal smears. Results: Compared to males (n=6), intravenous administration of pramlintide (6 µg/kg) significantly augmented the ongoing spontaneous activity and dural-evoked neuronal responses in the trigeminocervical complex, only during the estrus and early diestrus phases of the female estrous cycle, whereas this effect was not observed in the metestrus, proestrus and late diestrus phases (n=4-6 per group). Moreover, compared to vehicle (0.9% NaCl, n=5), intravenous administration of AC187 (6 µg/kg) significantly decreased the ongoing spontaneous and dural-evoked firing rates of central trigeminovascular neurons in males (n=4). Conclusion: Our data support that activation of the AMY1 receptor modulates the trigeminal nociceptive system and that this effect is most pronounced during the estrus and early diestrus phases of the mesntrual cycle. The data also support selective AMY1 antagonists as novel and potentially effective targets for the treatment of migraine.

Basic science, animal models in headache research

Neuropeptides, channels in headache

Abstract lecture

Prof. Dr.

Differential mechanism of action of erenumab and gepants in human isolated coronary arteries

Abstract text (incl. figure legends and references) Objective Multiple drugs targeting the calcitonin gene-related peptide (CGRP) pathway have been developed for the acute and preventive treatment of migraine. In this study, the effect of the monoclonal antibody erenumab in combination with the acute anti-migraine medication rimegepant, olcegepant or sumatriptan (5-HT-R agonist), on CGRP-induced vasorelaxation was investigated in human isolated coronary arteries (HCA). Methods HCA segments from 17 donors (11 female and 6 male, 53±3 years) were incubated overnight with 3 µM erenumab, which is the concentration of erenumab causing a maximum rightward shift of relaxations to CGRP. Next, the segments were mounted on a Mulvany myograph system, in the presence of 3 µM erenumab, and precontracted with 30 mM KCl and subsequently exposed to CGRP. Rimegepant, olcegepant or sumatriptan was added in increasing concentrations to assess whether these compounds did exert additional CGRP-blocking effects on top of erenumab. In addition, full concentration-response curves to CGRP, adrenomedullin or pramlintide were constructed in HCA segments incubated with or without erenumab (3 µM) and/or olcegepant (1 µM). Results The relaxation caused by CGRP in segments incubated with 3 µM erenumab was reversed by rimegepant (91%), olcegepant (92%) and sumatriptan (134%, physiological antagonism). Olcegepant seemed to further shift the concentration-response curve to CGRP when administered on top of erenumab, while no further shift of the responses to adrenomedullin and pramlintide was observed. Conclusion Both rimegepant and olcegepant exert additional effects on top of a maximal effect of erenumab, suggesting that the mechanism of action of erenumab and gepants may not be identical. In contrast, olcegepant does not induce additional effects on top of erenumab for the agonists adrenomedullin and pramlintide. This suggests that different receptor populations may mediate responses to CGRP, adrenomedullin and pramlintide in HCA.

Migraine

Revisiting RAMP1 expression: Characterisation of antibodies against receptor activity-modifying protein 1 (RAMP1)

Abstract text (incl. figure legends and references) Objective: Calcitonin gene-related peptide (CGRP) is well-established as a key component of migraine pathophysiology, yielding effective migraine therapeutics. Further refinement of these treatments requires a more in-depth understanding of CGRP's actions at its receptors, which contain a core accessory protein subunit: receptor activity-modifying protein 1 (RAMP1). Our understanding of RAMP1 expression at the cellular level is incomplete, partly due to the challenges in the availability and identification of specific and validated antibody tools. Methods: We profiled antibodies for immunodetection of RAMP1 using western blotting, immunocytochemistry, and immunohistochemistry, including using RAMP1 knockout mouse tissue. Results: Most antibodies could detect RAMP1 protein in western blotting and immunocytochemistry using transfected cells. Selected antibodies were profiled further, and two antibodies (844, ab256575) could detect a RAMP1-like band in western blots of rodent brain but not RAMP1 knockout mice. However, cross-reactivity with other proteins was evident for all antibodies. In immunohistochemistry clear conclusions about RAMP1 anatomical localization were prevented because each antibody unexpectedly detected distinct patterns of immunoreactivity. Conclusions: Although most antibodies were capable of detecting RAMP1, their ability to also detect off-target proteins means that immunoreactivity produced by RAMP1 antibodies (including 844) in nervous tissue cannot be confidently attributed to RAMP1 in immunohistochemical applications. RAMP1 expression in nervous tissue therefore needs to be revisited. Our results have implications for using RAMP1 antibodies for any immunohistochemical investigation in any tissue. Furthermore, as RAMP1 is important for other GPCR/ligand pairings, our results have broader significance beyond the CGRP field.

SciSe 5
Amylin physiology, receptors and potential new player in migraine

Appointment

Chair

Programme

SciSe 5Amylin physiology, receptors and potential new player in migraine

Appointment

Chair

Programme

SciSe 5
Amylin physiology, receptors and potential new player in migraine