| 1. |
- Edvinsson, Jacob C.A., et al.
(författare)
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C-fibers may modulate adjacent Aδ-fibers through axon-axon CGRP signaling at nodes of Ranvier in the trigeminal system
- 2019
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Ingår i: Journal of Headache and Pain. - : Springer Science and Business Media LLC. - 1129-2369 .- 1129-2377. ; 20:1
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Monoclonal antibodies (mAbs) towards CGRP or the CGRP receptor show good prophylactic antimigraine efficacy. However, their site of action is still elusive. Due to lack of passage of mAbs across the blood-brain barrier the trigeminal system has been suggested a possible site of action because it lacks blood-brain barrier and hence is available to circulating molecules. The trigeminal ganglion (TG) harbors two types of neurons; half of which store CGRP and the rest that express CGRP receptor elements (CLR/RAMP1). METHODS: With specific immunohistochemistry methods, we demonstrated the localization of CGRP, CLR, RAMP1, and their locations related to expression of the paranodal marker contactin-associated protein 1 (CASPR). Furthermore, we studied functional CGRP release separately from the neuron soma and the part with only nerve fibers of the trigeminal ganglion, using an enzyme-linked immunosorbent assay. RESULTS: Antibodies towards CGRP and CLR/RAMP1 bind to two different populations of neurons in the TG and are found in the C- and the myelinated Aδ-fibers, respectively, within the dura mater and in trigeminal ganglion (TG). CASPR staining revealed paranodal areas of the different myelinated fibers inhabiting the TG and dura mater. Double immunostaining with CASPR and RAMP1 or the functional CGRP receptor antibody (AA58) revealed co-localization of the two peptides in the paranodal region which suggests the presence of the CGRP-receptor. Double immunostaining with CGRP and CASPR revealed that thin C-fibers have CGRP-positive boutons which often localize in close proximity to the nodal areas of the CGRP-receptor positive Aδ-fibers. These boutons are pearl-like synaptic structures, and we show CGRP release from fibers dissociated from their neuronal bodies. In addition, we found that adjacent to the CGRP receptor localization in the node of Ranvier there was PKA immunoreactivity (kinase stimulated by cAMP), providing structural possibility to modify conduction activity within the Aδ-fibers. CONCLUSION: We observed a close relationship between the CGRP containing C-fibers and the Aδ-fibers containing the CGRP-receptor elements, suggesting a point of axon-axon interaction for the released CGRP and a site of action for gepants and the novel mAbs to alleviate migraine.
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| 2. |
- Edvinsson, Jacob Carl Alexander, et al.
(författare)
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Differences in pituitary adenylate cyclase-activating peptide and calcitonin gene-related peptide release in the trigeminovascular system
- 2020
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Ingår i: Cephalalgia. - : SAGE Publications. - 0333-1024 .- 1468-2982. ; 40:12, s. 1296-1309
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Tidskriftsartikel (refereegranskat)abstract
- Background: Several neurotransmitters are expressed in the neurons of the trigeminal ganglion. One such signalling molecule is the pituitary adenylate cyclase-activating peptide (PACAP). PACAP signalling has been suggested to have a possible role in the pathophysiology of primary headaches. Objective: The present study was designed to investigate the relationship between PACAP and calcitonin gene-related peptide, currently the two most relevant migraine peptides. Methods: In the current study, we used ELISA to investigate PACAP and calcitonin gene-related peptide release in response to 60 mM K+ or capsaicin using a rat hemi-skull model. We combined this analysis with qPCR and immunohistochemistry to study the expression of PACAP and calcitonin gene-related peptide receptors and ligands. Results: Calcitonin gene-related peptide (CGRP) is released from the trigeminal ganglion and dura mater. In contrast, PACAP is only released from the trigeminal ganglion. We observed a weak correlation between the stimulated release of the two neuropeptides. PACAP-38 immunoreactivity was expressed alone and in a subpopulation of neurons in the trigeminal ganglion that also store calcitonin gene-related peptide. The receptor subtype PAC1 was mainly expressed in the satellite glial cells (SGCs), which envelop the neurons in the trigeminal ganglion, in some neuronal processes, inside the Aδ-fibres and in the outermost layer of the myelin sheath that envelopes the Aδ-fibres. Conclusion: Unlike CGRP, PACAP is only released within the trigeminal ganglion. This raises the question of whether a migraine therapy aimed at preventing peripheral PACAP signalling would be as successful as the CGRP signalling targeted treatments.
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| 3. |
- Edvinsson, Jacob C.A., et al.
(författare)
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Neurokinins and their receptors in the rat trigeminal system : Differential localization and release with implications for migraine pain
- 2021
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Ingår i: Molecular Pain. - : SAGE Publications. - 1744-8069. ; 17
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Tidskriftsartikel (refereegranskat)abstract
- Substance P (SP) and calcitonin gene-related peptide (CGRP) have both been considered potential drug candidates in migraine therapy. In recent years, CGRP receptor inhibition has been established as an effective treatment, in particular as a prophylactic for chronic migraine. Curiously, inhibition of neurokinin receptor 1 (NK1R) failed to alleviate acute migraine attacks in clinical trials, and the neurokinins were consequently abandoned as potential antimigraine candidates. The reason behind this has remained enigmatic. Utilizing immunohistochemistry and semi-quantitative cell counts the expression of neurokinins and their associated receptors was examined in the rat trigeminal ganglion. Immunohistochemistry results revealed SP co-localization in CGRP positive neurons and C-fibres, where it mainly concentrated at boutons. Neurokinin A (NKA) was observed in a population of C-fibres and small neurons where it could co-localize with SP. In contrast, neurokinin B (NKB) did not co-localize with SP and was observed in large/medium sized neurons and Aδ-fibres. All neurokinin receptors (NK1-3R) were found to be expressed in a majority of trigeminal ganglion neurons and A-fibres. The functional release of SP and CGRP in the trigeminovascular system was stimulated with either 60 mM K+ or 100 nM capsaicin and measured with an enzyme-linked immunosorbent assay (ELISA). ELISA results established that SP can be released locally from trigeminovascular system. The released SP was comparatively minor compared to the CGRP release from stimulated dura mater, trigeminal ganglion neurons and fibres. We hypothesize that SP and CGRP signalling pathways may work in tandem to exacerbate painful stimuli in the TGV system.
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| 4. |
- Erdling, André, et al.
(författare)
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Differential inhibitory response to telcagepant on αCGRP induced vasorelaxation and intracellular Ca2+ levels in the perfused and non-perfused isolated rat middle cerebral artery
- 2017
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Ingår i: Journal of Headache and Pain. - : Springer Science and Business Media LLC. - 1129-2369 .- 1129-2377. ; 18:1, s. 1-9
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Tidskriftsartikel (refereegranskat)abstract
- Background: Calcitonin gene-related peptide (CGRP) is one of the most potent endogenous vasodilators identified to date. The present study elucidates the differential interaction of CGRP, its receptor and the effect of the CGRP-receptor antagonist telcagepant on intracellular Ca2+ -levels and tension in rat middle cerebral arteries (MCA) by pressurized arteriography, FURA-2/wire myography and immunohistochemistry. Methods: A pressurized arteriograph system was used to evaluate changes in MCA tension when subjected to CGRP and/or telcagepant. Intracellular calcium levels were evaluated using a FURA-2/wire myograph system. Localization of the CGRP-receptor components was verified using immunohistochemistry. Results: Abluminal but not luminal αCGRP (10-12-10-6 M) caused concentration-dependent vasorelaxation in rat MCA. Luminal telcagepant (10-6 M) failed to inhibit this relaxation, while abluminal telcagepant inhibited the relaxation (10-6 M). Using the FURA-2 method in combination with wire myography we observed that αCGRP reduced intracellular calcium levels and in parallel the vascular tone. Telcagepant (10-6 M) inhibited both vasorelaxation and drop in intracellular calcium levels. Both functional components of the CGRP receptor, CLR (calcitonin receptor-like receptor) and RAMP1 (receptor activity modifying peptide 1) were found in the smooth muscle cells but not in the endothelial cells of the cerebral vasculature. Conclusions: This study thus demonstrates the relaxant effect of αCGRP on rat MCA. The vasorelaxation is associated with a simultaneous decrease in intracellular calcium levels. Telcagepant reduced relaxation and thwarted the reduction in intracellular calcium levels localized in the vascular smooth muscle cells. In addition, telcagepant may act as a non-competitive antagonist at concentrations greater than 10-8 M.
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| 5. |
- Erdling, André, et al.
(författare)
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VIP/PACAP receptors in cerebral arteries of rat: Characterization, localization and relation to intracellular calcium.
- 2013
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Ingår i: Neuropeptides. - : Elsevier BV. - 1532-2785 .- 0143-4179. ; 47:2, s. 85-92
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase activating peptide (PACAP)-containing nerves surround cerebral blood vessels. The peptides have potent vasodilator properties via smooth muscle cell receptors and activation of adenylate cyclase. The purpose of this study was to describe the effects of two putative VIP/PACAP receptor antagonists and the distribution of the receptor protein in rat brain vessels. METHODS: The vascular effects of VIP, PACAP-27 and PACAP-38 were investigated in segments of rat middle cerebral artery (MCA) by pressurized arteriography, and in a wire myograph. The antagonistic responses to PACAP6-38 and PG99-465 were evaluated. In addition, the receptor subtypes for VIP and PACAP (VPAC(1), VPAC(2) and PAC(1)) were visualized in the rat middle cerebral artery by immunohistochemistry and Western blotting. RESULTS: In the perfusion model, abluminal but not luminal VIP, PACAP-27 and PACAP-38 caused concentration-dependent relaxations of the MCA (27.1±0.2%, 25.2±0.4% and 0.3±0.1%, respectively). In the wire myograph, there was no significant difference in potency of the peptides in the MCA. In both systems, PACAP6-38 and PG99-465 inhibited the VIP induced relaxation. Western blot showed the presence of the receptor proteins in cerebral vasculature and immunohistochemistry showed that all three receptors are present and located in the cytoplasm of smooth muscle cells. CONCLUSION: In both systems, the two blockers antagonized the relaxant VIP effect; the potency order of agonists and the immunohistochemistry suggest the presence of the dilatory VPAC(1) and VPAC(2) receptors on the smooth muscle cells.
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| 6. |
- Johansson, Sara Ellinor, et al.
(författare)
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Vascular pathology of large cerebral arteries in experimental subarachnoid hemorrhage : Vasoconstriction, functional CGRP depletion and maintained CGRP sensitivity
- 2019
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Ingår i: European Journal of Pharmacology. - : Elsevier BV. - 0014-2999. ; 846, s. 109-118
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Tidskriftsartikel (refereegranskat)abstract
- Subarachnoid hemorrhage (SAH) is associated with increased cerebral artery sensitivity to vasoconstrictors and release of the perivascular sensory vasodilator CGRP. In the current study the constrictive phenotype and the vasodilatory effects of exogenous and endogenous perivascular CGRP were characterized in detail applying myograph technology to cerebral artery segments isolated from experimental SAH and sham-operated rats. Following experimental SAH, cerebral arteries exhibited increased vasoconstriction to endothelin-1, 5-hydroxytryptamine and U46419. In addition, depolarization-induced vasoconstriction (60 mM potassium) was significantly increased, supporting a general SAH-associated vasoconstrictive phenotype. Using exogenous CGRP, we demonstrated that sensitivity of the arteries to CGRP-induced vasodilation was unchanged after SAH. However, vasodilation in response to capsaicin (100 nM), a sensory nerve activator used to release perivascular CGRP, was significantly reduced by SAH (P = 0.0079). Because CGRP-mediated dilation is an important counterbalance to increased arterial contractility, a reduction in CGRP release after SAH would exacerbate the vasospasms that occur after SAH. A similar finding was obtained with artery culture (24 h), an in vitro model of SAH-induced vascular dysfunction. The arterial segments maintained sensitivity to exogenous CGRP but showed reduced capsaicin-induced vasodilation. To test whether a metabolically stable CGRP analogue could be used to supplement the loss of perivascular CGRP release in SAH, SAX was systemically administered in our in vivo SAH model. SAX treatment, however, induced CGRP-desensitization and did not prevent the development of vasoconstriction in cerebral arteries after SAH.
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| 7. |
- Le, Thi Lisa, et al.
(författare)
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CGRP in rat mesenteric artery and vein - receptor expression, CGRP presence and potential roles
- 2020
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Ingår i: European Journal of Pharmacology. - : Elsevier BV. - 0014-2999. ; 875
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Tidskriftsartikel (refereegranskat)abstract
- CGRP is a potent dilator of arteries and despite rich perivascular CGRP immunoreactivity in both arteries and veins the role of CGRP in veins remains unknown. The aim of the current study was to compare perivascular CGRP immunoreactivity and expression of CGRP receptor mRNA and CGRP receptor immunoreactivity in rat mesenteric arteries and veins. Furthermore, potential vasomotor effects of CGRP were explored in veins. Immunohistochemical studies reproduced rich perivascular CGRP innervation in arteries and in veins. Further, the presence of mRNA encoding the CGRP receptor subunits, CLR and RAMP1, were demonstrated in both arteries and veins using qPCR. Before comparing the vasoactive effects of CGRP in arteries and veins, we aimed to identify an experimental setting where vasomotor responses could be detected. Therefore, a length-tension study was performed in artery and vein segments. Whereas the arteries showed the characteristic monophasic curve with an IC/IC100 value of 0.9, surprisingly the veins showed a biphasic response with two corresponding IC/IC100 values of 0.7 and 0.9, respectively. There was no significant difference between fresh and cultured vasculature segments. To investigate whether a potential tension-dependent CGRP-induced dilation of veins caused the decline between the two IC/IC100 peaks, a second study was performed, with the CGRP receptor antagonist, BIBN4096BS (olcegepant) and the sensory nerve secretagogue, capsaicin. No significant vascular role of endogenous perivascular CGRP in mesenteric veins could be concluded, and a potential role of the rich perivascular CGRP and CGRP receptor abundancy in veins remains unknown.
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| 8. |
- Rasmussen, Marianne N. P., et al.
(författare)
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Permanent Distal Occlusion of Middle Cerebral Artery in Rat Causes Local Increased ETB, 5-HT1B and AT(1) Receptor-Mediated Contractility Downstream of Occlusion
- 2013
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Ingår i: Journal of Vascular Research. - : S. Karger AG. - 1423-0135 .- 1018-1172. ; 50:5, s. 396-409
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Tidskriftsartikel (refereegranskat)abstract
- Background/Aims: In response to experimental stroke, a characteristic functional and expressional upregulation of contractile G-protein-coupled receptors has been uncovered in the affected cerebral vasculature; however, the mechanism initiating this phenomenon remains unknown. Methods: Using a model of permanent distal occlusion of rat middle cerebral arteries, we investigated whether there was a regional difference in receptor-mediated contractility of segments located upstream and downstream of the occlusion site. The contractile response to endothelin, angiotensin and 5-hydroxytryptamine receptor stimulation was studied by sensitive wire myograph. Results: Only downstream segments exhibited an augmented contractile response to stimulation with each of the three ligands, with the response towards sarafotoxin 6c being especially augmented compared to sham, upstream and contralateral controls. This functional increase did not seem to relate to ischemic tissue damage, inflammatory cell infiltration or the element of reperfusion. Interestingly, immunohistochemistry did not show any difference in the level of immunoreactivity towards endothelin B (ETB) receptors between groups. Conclusion: Single artery occlusion without significant visible infarct resulted in locally increased ETB, angiotensin type 1 and 5-hydroxytryptamine 1B receptor-mediated contractile responses only in segments located downstream of the occlusion site. This suggests lack of wall stress as an initiating trigger leading to regulation of contractile response after cerebral stroke. (C) 2013 S. Karger AG, Basel
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| 9. |
- Sohn, Iben, et al.
(författare)
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The effects of CGRP in vascular tissue - Classical vasodilation, shadowed effects and systemic dilemmas
- 2020
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Ingår i: European Journal of Pharmacology. - : Elsevier BV. - 0014-2999. ; 881
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Forskningsöversikt (refereegranskat)abstract
- Vascular tissue consists of endothelial cells, vasoactive smooth muscle cells and perivascular nerves. The perivascular sensory neuropeptide CGRP has demonstrated potent vasodilatory effects in any arterial vasculature examined so far, and a local protective CGRP-circuit of sensory nerve terminal CGRP release and smooth muscle cell CGRP action is evident. The significant vasodilatory effect has shadowed multiple other effects of CGRP in the vascular tissue and we therefore thoroughly review vascular actions of CGRP on endothelial cells, vascular smooth muscle cells and perivascular nerve terminals. The actions beyond vasodilation includes neuronal re-uptake and neuromodulation, angiogenic, proliferative and antiproliferative, pro- and anti-inflammatory actions which vary depending on the target cell and anatomical location. In addition to the classical perivascular nerve-smooth muscle CGRP circuit, we review existing evidence for a shadowed endothelial autocrine pathway for CGRP. Finally, we discuss the impact of local and systemic actions of CGRP in vascular regulation and protection from hypertensive and ischemic heart conditions with special focus on therapeutic CGRP agonists and antagonists.
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| 10. |
- Spray, Stine, et al.
(författare)
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Reduced Mechanical Stretch Induces Enhanced Endothelin B Receptor-mediated Contractility via Activation of Focal Adhesion Kinase and Extra Cellular-regulated Kinase 1/2 in Cerebral Arteries from Rat.
- 2016
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Ingår i: Basic & Clinical Pharmacology & Toxicology. - : Wiley. - 1742-7843 .- 1742-7835. ; 119:1, s. 68-77
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Tidskriftsartikel (refereegranskat)abstract
- Cerebral ischaemia results in enhanced endothelin B (ETB ) receptor-mediated contraction and receptor protein expression in the affected cerebrovascular smooth muscle cells (SMC). Organ culture of cerebral arteries is a method to induce similar alterations in ETB receptor expression. We hypothesize that rapid and sustained reduction in wall tension/stretch is a possible trigger mechanism for this vascular remodelling. Isolated rat middle cerebral artery (MCA) segments were incubated in a wire-myograph with or without mechanical stretch, prior to assessment of their contractile response to the selective ETB receptor agonist sarafotoxin 6c. The involvement of extracellular regulated kinase (ERK) 1/2 and focal adhesion kinase (FAK) were studied by their specific inhibitors U0126 and PF-228, respectively. Compared to their stretched counterparts, un-stretched MCA segments showed a significantly increased ETB receptor-mediated contractile response following 12 hr of incubation, which was attenuated by either U0126 or PF-228. The functionally increased ETB -mediated contractility could be attributed to two different mechanisms: 1) a difference in ETB receptor localization from primarily endothelial expression to SMC expression and 2) an increased calcium sensitivity of the SMCs due to an increased expression of the calcium channel transient receptor potential canonical 1. Collectively, our results present a possible mechanism linking lack of vessel wall stretch/tension to changes in ETB receptor-mediated contractility via triggering of an early mechanosensitive signalling pathway involving ERK1/2 and FAK signalling. A mechanism likely to be an initiating factor for the increased ETB receptor-mediated contractility found after cerebral ischaemia. This article is protected by copyright. All rights reserved.
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