Cluster headache and kynurenines.

BACKGROUND: The glutamatergic neurotransmission has important role in the pathomechanism of primary headache disorders. The kynurenine metabolites derived from catabolism of tryptophan (Trp) have significant involvement not only in glutamatergic processes, but also in the neuroinflammation, the oxidative stress and the mitochondrial dysfunctions. Previously we identified a depressed peripheral Trp metabolism in interictal period of episodic migraineurs, which prompted us to examine this pathway in patients with episodic cluster headache (CH) as well. Our aims were to compare the concentrations of compounds both in headache-free and attack periods, and to find correlations between Trp metabolism and the clinical features of CH. Levels of 11 molecules were determined in peripheral blood plasma of healthy controls (n = 22) and interbout/ictal periods of CH patients (n = 24) by neurochemical measurements. FINDINGS: Significantly decreased L-kynurenine (KYN, p < 0.01), while increased quinolinic acid (QUINA, p < 0.005) plasma concentrations were detected in the interbout period of CH patients compared to healthy subjects. The levels of KYN are further reduced during the ictal period compared to the controls (p < 0.006). There was a moderate, negative correlation between disease duration and interbout QUINA levels (p < 0.048, R = - 0.459); and between the total number of CH attacks experienced during the lifetime of patients and the interbout KYN concentrations (p < 0.024, R = - 0.516). Linear regression models revealed negative associations between age and levels of Trp, kynurenic acid, 3-hdyroxyanthranilic acid and QUINA in healthy control subjects, as well as between age and ictal level of anthranilic acid. CONCLUSIONS: Our results refer to a specifically altered Trp metabolism in CH patients. The onset of metabolic imbalance can be attributed to the interbout period, where the decreased KYN level is unable to perform its protective functions, while the concentration of QUINA, as a toxic compound, increases. These processes can trigger CH attacks, which may be associated with glutamate excess induced neurotoxicity, neuroinflammation and oxidative stress. Further studies are needed to elucidate the exact functions of these molecular alterations that can contribute to identify new, potential biomarkers in the therapy of CH.

Disease- and headache-specific microRNA signatures and their predicted mRNA targets in peripheral blood mononuclear cells in migraineurs: role of inflammatory signalling and oxidative stress.

BACKGROUND: Migraine is a primary headache with genetic susceptibility, but the pathophysiological mechanisms are poorly understood, and it remains an unmet medical need. Earlier we demonstrated significant differences in the transcriptome of migraineurs' PBMCs (peripheral blood mononuclear cells), suggesting the role of neuroinflammation and mitochondrial dysfunctions. Post-transcriptional gene expression is regulated by miRNA (microRNA), a group of short non-coding RNAs that are emerging biomarkers, drug targets, or drugs. MiRNAs are emerging biomarkers and therapeutics; however, little is known about the miRNA transcriptome in migraine, and a systematic comparative analysis has not been performed so far in migraine patients. METHODS: We determined miRNA expression of migraineurs' PBMC during (ictal) and between (interictal) headaches compared to age- and sex-matched healthy volunteers. Small RNA sequencing was performed from the PBMC, and mRNA targets of miRNAs were predicted using a network theoretical approach by miRNAtarget.com™. Predicted miRNA targets were investigated by Gene Ontology enrichment analysis and validated by comparing network metrics to differentially expressed mRNA data. RESULTS: In the interictal PBMC samples 31 miRNAs were differentially expressed (DE) in comparison to healthy controls, including hsa-miR-5189-3p, hsa-miR-96-5p, hsa-miR-3613-5p, hsa-miR-99a-3p, hsa-miR-542-3p. During headache attacks, the top DE miRNAs as compared to the self-control samples in the interictal phase were hsa-miR-3202, hsa-miR-7855-5p, hsa-miR-6770-3p, hsa-miR-1538, and hsa-miR-409-5p. MiRNA-mRNA target prediction and pathway analysis indicated several mRNAs related to immune and inflammatory responses (toll-like receptor and cytokine receptor signalling), neuroinflammation and oxidative stress, also confirmed by mRNA transcriptomics. CONCLUSIONS: We provide here the first evidence for disease- and headache-specific miRNA signatures in the PBMC of migraineurs, which might help to identify novel targets for both prophylaxis and attack therapy.

Memory Enhancement with Kynurenic Acid and Its Mechanisms in Neurotransmission.

Kynurenic acid (KYNA) is an endogenous tryptophan (Trp) metabolite known to possess neuroprotective property. KYNA plays critical roles in nociception, neurodegeneration, and neuroinflammation. A lower level of KYNA is observed in patients with neurodegenerative diseases such as Alzheimer's and Parkinson's diseases or psychiatric disorders such as depression and autism spectrum disorders, whereas a higher level of KYNA is associated with the pathogenesis of schizophrenia. Little is known about the optimal concentration for neuroprotection and the threshold for neurotoxicity. In this study the effects of KYNA on memory functions were investigated by passive avoidance test in mice. Six different doses of KYNA were administered intracerebroventricularly to previously trained CFLP mice and they were observed for 24 h. High doses of KYNA (i.e., 20-40 µg/2 µL) significantly decreased the avoidance latency, whereas a low dose of KYNA (0.5 µg/2 µL) significantly elevated it compared with controls, suggesting that the low dose of KYNA enhanced memory function. Furthermore, six different receptor blockers were applied to reveal the mechanisms underlying the memory enhancement induced by KYNA. The series of tests revealed the possible involvement of the serotonergic, dopaminergic, α and ß adrenergic, and opiate systems in the nootropic effect. This study confirmed that a low dose of KYNA improved a memory component of cognitive domain, which was mediated by, at least in part, four systems of neurotransmission in an animal model of learning and memory.

Identification of disease- and headache-specific mediators and pathways in migraine using blood transcriptomic and metabolomic analysis.

BACKGROUND: Recent data suggest that gene expression profiles of peripheral white blood cells can reflect changes in the brain. We aimed to analyze the transcriptome of peripheral blood mononuclear cells (PBMC) and changes of plasma metabolite levels of migraineurs in a self-controlled manner during and between attacks. METHODS: Twenty-four patients with migraine were recruited and blood samples were collected in a headache-free (interictal) period and during headache (ictal) to investigate disease- and headache-specific alterations. Control samples were collected from 13 age- and sex-matched healthy volunteers. RNA was isolated from PBMCs and single-end 75 bp RNA sequencing was performed using Illumina NextSeq 550 instrument followed by gene-level differential expression analysis. Functional analysis was carried out on information related to the role of genes, such as signaling pathways and biological processes. Plasma metabolomic measurement was performed with the Biocrates MxP Quant 500 Kit. RESULTS: We identified 144 differentially-expressed genes in PBMCs between headache and headache-free samples and 163 between symptom-free patients and controls. Network analysis revealed that enriched pathways included inflammation, cytokine activity and mitochondrial dysfunction in both headache and headache-free samples compared to controls. Plasma lactate, succinate and methionine sulfoxide levels were higher in migraineurs while spermine, spermidine and aconitate were decreased during attacks. CONCLUSIONS: It is concluded that enhanced inflammatory and immune cell activity, and oxidative stress can play a role in migraine susceptibility and headache generation.

Clinical relevance of depressed kynurenine pathway in episodic migraine patients: potential prognostic markers in the peripheral plasma during the interictal period.

BACKGROUND: Altered glutamatergic neurotransmission and neuropeptide levels play a central role in migraine pathomechanism. Previously, we confirmed that kynurenic acid, an endogenous glutamatergic antagonist, was able to decrease the expression of pituitary adenylate cyclase-activating polypeptide 1-38, a neuropeptide with known migraine-inducing properties. Hence, our aim was to reveal the role of the peripheral kynurenine pathway (KP) in episodic migraineurs. We focused on the complete tryptophan (Trp) catabolism, which comprises the serotonin and melatonin routes in addition to kynurenine metabolites. We investigated the relationship between metabolic alterations and clinical characteristics of migraine patients. METHODS: Female migraine patients aged between 25 and 50 years (n = 50) and healthy control subjects (n = 34) participated in this study. Blood samples were collected from the cubital veins of subjects (during both the interictal/ictal periods in migraineurs, n = 47/12, respectively). 12 metabolites of Trp pathway were determined by neurochemical measurements (UHPLC-MS/MS). RESULTS: Plasma concentrations of the most Trp metabolites were remarkably decreased in the interictal period of migraineurs compared to healthy control subjects, especially in the migraine without aura (MWoA) subgroup: Trp (p < 0.025), L-kynurenine (p < 0.001), kynurenic acid (p < 0.016), anthranilic acid (p < 0.007), picolinic acid (p < 0.03), 5-hydroxy-indoleaceticacid (p < 0.025) and melatonin (p < 0.023). Several metabolites showed a tendency to elevate during the ictal phase, but this was significant only in the cases of anthranilic acid, 5-hydroxy-indoleaceticacid and melatonin in MWoA patients. In the same subgroup, higher interictal kynurenic acid levels were identified in patients whose headache was severe and not related to their menstruation cycle. Negative linear correlation was detected between the interictal levels of xanthurenic acid/melatonin and attack frequency. Positive associations were found between the ictal 3-hydroxykynurenine levels and the beginning of attacks, just as between ictal picolinic acid levels and last attack before ictal sampling. CONCLUSIONS: Our results suggest that there is a widespread metabolic imbalance in migraineurs, which manifests in a completely depressed peripheral Trp catabolism during the interictal period. It might act as trigger for the migraine attack, contributing to glutamate excess induced neurotoxicity and generalised hyperexcitability. This data can draw attention to the clinical relevance of KP in migraine.

Temporal instability of salience network activity in migraine with aura.

This study aims to investigate whether intranetwork dynamic functional connectivity and causal interactions of the salience network is altered in the interictal term of migraine. Thirty-two healthy controls, 37 migraineurs without aura, and 20 migraineurs with aura were recruited. Participants underwent a T1-weighted scan and resting-state fMRI protocol inside a 1.5T MR scanner. We obtained average spatial maps of resting-state networks using group independent component analysis, which yielded subject-specific time series through a dual regression approach. Salience network regions of interest (bilateral insulae and prefrontal cortices, dorsal anterior cingulate cortex) were obtained from the group average map through cluster-based thresholding. To describe intranetwork connectivity, average and dynamic conditional correlation was calculated. Causal interactions between the default-mode, dorsal attention, and salience network were characterised by spectral Granger's causality. Time-averaged correlation was lower between the right insula and prefrontal cortex in migraine without aura vs with aura and healthy controls (P < 0.038, P < 0.037). Variance of dynamic conditional correlation was higher in migraine with aura vs healthy controls and migraine with aura vs without aura between the right insula and dorsal anterior cingulate cortex (P < 0.011, P < 0.026), and in migraine with aura vs healthy controls between the dorsal anterior cingulate and left prefrontal cortex (P < 0.021). Causality was weaker in the <0.05 Hz frequency range between the salience and dorsal attention networks in migraine with aura (P < 0.032). Overall, migraineurs with aura exhibit more fluctuating connections in the salience network, which also affect network interactions, and could be connected to altered cortical excitability and increased sensory gain.

Are Migraine With and Without Aura Really Different Entities?

Background: Migraine research is booming with the rapidly developing neuroimaging tools. Structural and functional alterations of the migrainous brain were detected with MRI. The outcome of a research study largely depends on the working hypothesis, on the chosen measurement approach and also on the subject selection. Against all evidence from the literature that migraine subtypes are different, most of the studies handle migraine with and without aura as one disease. Methods: Publications from PubMed database were searched for terms of "migraine with aura," "migraine without aura," "interictal," "MRI," "diffusion weighted MRI," "functional MRI," "compared to," "atrophy" alone and in combination. Conclusion: Only a few imaging studies compared the two subforms of the disease, migraine with aura, and without aura, directly. Functional imaging investigations largely agree that there is an increased activity/activation of the brain in migraine with aura as compared to migraine without aura. We propose that this might be the signature of cortical hyperexcitability. However, structural investigations are not equivocal. We propose that variable contribution of parallel, competing mechanisms of maladaptive plasticity and neurodegeneration might be the reason behind the variable results.

The effect of orofacial complete Freund's adjuvant treatment on the expression of migraine-related molecules.

BACKGROUND: Migraine is a neurovascular primary headache disorder, which causes significant socioeconomic problems worldwide. The pathomechanism of disease is enigmatic, but activation of the trigeminovascular system (TS) appears to be essential during the attack. Migraine research of recent years has focused on neuropeptides, such as calcitonin gene-related peptide (CGRP) and pituitary adenylate cyclase-activating polypeptide 1-38 (PACAP1-38) as potential pathogenic factors and possible therapeutic offensives. The goal of present study was to investigate the simultaneous expression of CGRP and precursor of PACAP1-38 (preproPACAP) in the central region of the TS in a time-dependent manner following TS activation in rats. METHODS: The right whisker pad of rats was injected with 50 µl Complete Freund's Adjuvant (CFA) or saline. A mechanical allodynia test was performed with von Frey filaments before and after treatment. Transcardial perfusion of the animals was initiated 24, 48, 72 and 120 h after injection, followed by the dissection of the nucleus trigeminus caudalis (TNC). After preparation, the samples were stored at - 80 °C until further use. The relative optical density of CGRP and preproPACAP was analyzed by Western blot. One-way ANOVA and Kruskal-Wallis followed by Tukey post hoc test were used to evaluate the data. Regression analysis was applied to explore the correlation between neuropeptides expression and hyperalgesia. RESULTS: Orofacial CFA injection resulted in significant CGRP and preproPACAP release in the TNC 24, 48, 72 and 120 h after the treatment. The level of neuropeptides reached its maximum at 72 h after CFA injection, corresponding to the peak of facial allodynia. Negative, linear correlation was detected between the expression level of neuropeptides and value of mechanonociceptive threshold. CONCLUSION: This is the first study which suggests that the expression of CGRP and preproPACAP simultaneously increases in the central region of activated TS and it influences the formation of mechanical hyperalgesia. Our results contribute to a better understanding of migraine pathogenesis and thereby to the development of more effective therapeutic approaches.

Correlation of neurochemical and imaging markers in migraine: PACAP38 and DTI measures.

OBJECTIVE: To examine whether interictal plasma pituitary adenylate cyclase-activating peptide 38-like immunoreactivity (PACAP38-LI) shows correlation with the microstructural integrity of the white matter in migraine. METHODS: Interictal plasma PACAP38-LI was measured by radioimmunoassay in 26 patients with migraine (24 women) who underwent diffusion tensor imaging afterward using a 1.5-tesla magnetic resonance scanner. Data were analyzed using tract-based spatial statistics included in FMRIB's Software Library. RESULTS: Interictal plasma PACAP38-LI showed significant correlation with mean diffusivity (p < 0.0179) mostly in the bilateral occipital white matter spreading into parietal and temporal white matter. Axial and radial diffusivity showed positive correlation with interictal PACAP38-LI (p < 0.0432 and p < 0.0418, respectively) in the left optic radiation and left posterior corpus callosum. Fractional anisotropy did not correlate significantly with PACAP38-LI. With disease duration as a nuisance regressor in the model, PACAP38-LI correlated with axial and mean diffusivity in the left thalamus (p < 0.01). CONCLUSION: We report a link between PACAP38, a pathobiologically important neurochemical biomarker, and imaging markers of the disease that may bolster further research into the role of PACAP38 in migraine.

Kynurenic acid and its derivatives are able to modulate the adhesion and locomotion of brain endothelial cells.

The neuroprotective actions of kynurenic acid (KYNA) and its derivatives in several neurodegenerative disorders [characterized by damage to the cerebral endothelium and to the blood-brain barrier (BBB)] are well established. Cell-extracellular matrix (ECM) adhesion is supposedly involved in recovery of impaired cerebral endothelium integrity (endothelial repair). The present work aimed to investigate the effects of KYNA and its synthetic derivatives on cellular behaviour (e.g. adhesion and locomotion) and on morphology of the GP8 rat brain endothelial cell line, modeling the BBB endothelium. The effects of KYNA and its derivatives on cell adhesion were measured using an impedance-based technique, the xCELLigence SP system. Holographic microscopy (Holomonitor™ M4) was used to analyse both chemokinetic responses and morphometry. The GP8 cells proved to be a suitable model cell line for investigating cell adhesion and the locomotion modulator effects of kynurenines. KYNA enhanced cell adhesion and spreading, and also decreased the migration/motility of GP8 cells at physiological concentrations (10-9 and 10-7 mol/L). The derivatives containing an amide side-chain at the C2 position (KYNA-A1 and A2) had lower adhesion inducer effects compared to KYNA. All synthetic analogues (except KYNA-A5) had a time-dependent inhibitory effect on GP8 cell adhesion at a supraphysiological concentration (10-3 mol/L). The immobilization promoting effect of KYNA and the adhesion inducer activity of its derivatives indicate that these compounds could contribute to maintaining or restoring the protective function of brain endothelium; they also suggest that cell-ECM adhesion and related cell responses (e.g. migration/motility) could be potential new targets of KYNA.

Macro- and microstructural alterations of the subcortical structures in episodic cluster headache.

Background Previous functional and structural imaging studies have revealed that subcortical structures play a key a role in pain processing. The recurring painful episodes might trigger maladaptive plasticity or alternatively degenerative processes that might be detected by MRI as changes in size or microstructure. In the current investigation, we aimed to identify the macro- and microstructural alterations of the subcortical structures in episodic cluster headache. Methods High-resolution T1-weighted and diffusion-weighted MRI images with 60 gradient directions were acquired from 22 patients with cluster headache and 94 healthy controls. Surface-based segmentation analysis was used to measure the volume of the subcortical nuclei, and mean diffusion parameters (fractional anisotropy, mean, radial and axial diffusivity) were determined for these structures. In order to understand whether the size and diffusion parameters could be investigated in a headache lateralised manner, first the asymmetry of the size and diffusion parameters of the subcortical structures was analysed. Volumes and diffusion parameters were compared between groups and correlated with the cumulative number of headache days. To account for the different size of the patient and control group, a bootstrap approach was used to investigate the stability of the findings. Results A significant lateralisation of the size (caudate, putamen and thalamus) and the diffusion parameters of the subcortical structures were found in normal controls. In cluster headache patients, the mean fractional anisotropy of the right amygdalae, the mean axial and mean diffusivity of the right caudate nucleus and the radial diffusivity of the right pallidum were higher. The mean anisotropy of the right pallidum was lower in patients. Conclusion The analysis of the pathology in the subcortical structures in episodic cluster headache reveals important features of the disease, which might allow a deeper insight into the pathomechanism of the pain processing in this headache condition.

Kynurenic acid and its analogue can alter the opioid receptor G-protein signaling after acute treatment via NMDA receptor in rat cortex and striatum.

Previously, we have shown that the N-methyl d-aspartate (NMDA)-receptor antagonist kynurenic acid (KYNA) and its analogue KYNA1 do not bind directly to mu, kappa and delta opioid receptors in vitro. On the other hand, chronic administration of KYNA and KYNA1 resulted in region (cortex vs striatum) and opioid receptor-type specific alterations in G-protein activation of mouse brain homogenates. Here we describe for the first time the acute effect of KYNA and KYNA1 on opioid receptor function with the possible involvement of the NMDA receptor. The acute 30minute in vivo KYNA1 and KYNA treatments altered opioid receptor G-protein signaling or ligand potency depending on the opioid receptor type and brain region (rat cortex vs striatum) using [35S]GTPγS binding assays. Pretreatment with the NMDA receptor antagonist MK-801 impaired or reversed the effects of KYNA1 and KYNA. These results suggest an NMDA receptor mediated effect. After acute 30minute treatment HPLC measurements revealed a similar KYNA1 and a higher KYNA plasma concentration compared to cerebrospinal fluid concentrations. Finally, KYNA, KYNA1 and MK-801 showed comparable results in opioid receptor G-protein activity and ligand potency with acute in vivo treatments when they were administered in vitro for 30min on isolated cortex and striatum slices. We previously demonstrated that KYNA1 and KYNA acutely altered opioid receptor function in vivo and in vitro through the NMDA receptor depending on the opioid receptor type and brain region. This study may lead to a new, indirect approach to influence opioid receptor signaling.

Kynurenines and PACAP in Migraine: Medicinal Chemistry and Pathogenetic Aspects.

BACKGROUND: Migraine is a highly disabling neurovascular primary headache disorder, with its exact pathomechanism being still unrevealed. The current leading hypotheses are based on the sensitization and activation of the trigeminovascular system. OBJECTIVE: To review the literature with focus on the effects of kynurenines (L-kynurenine and kynurenic acid) and pituitary adenylate cyclase-activating polypeptide on the regulation of the trigeminovascular system. METHOD: A literature search was conducted to identify preclinical and clinical publications (198 references) by using the keywords 'kynurenines', 'pituitary adenylate cyclase-activating polypeptide', and 'migraine' in the database of MEDLINE/PubMed up to 10 September 2016 for topical review. Additional filters used included 'review', 'systematic review', 'original article', and 'English language'. RESULTS: L-kynurenine and kynurenic acid act on the glutamatergic system at the level of the second-order nociceptive neurons in the trigeminal nucleus caudalis. Pituitary adenylate cyclase- activating polypeptide is released from the peripheral nerve endings of the trigeminal pseudounipolar neurons and causes vasodilation and mast cell degranulation, leading to consequent peripheral sensitization of the dural nociceptors. Centrally released pituitary adenylate cyclase-activating polypeptide in the trigeminal nucleus caudalis results in the central sensitization of the second-order neurons. The sensitization process leads to the characteristic features of migraine. CONCLUSION: L-kynurenine, kynurenic acid, and pituitary adenylate cyclase-activating polypeptide may have fundamental roles in the initiation of migraine headache attacks.

Interictal brain activity differs in migraine with and without aura: resting state fMRI study.

BACKGROUND: Migraine is one of the most severe primary headache disorders. The nature of the headache and the associated symptoms during the attack suggest underlying functional alterations in the brain. In this study, we examined amplitude, the resting state fMRI fluctuation in migraineurs with and without aura (MWA, MWoA respectively) and healthy controls. METHODS: Resting state functional MRI images and T1 high-resolution images were acquired from all participants. For data analysis we compared the groups (MWA-Control, MWA-MWoA, MWoA-Control). The resting state networks were identified by MELODIC. The mean time courses of the networks were identified for each participant for all networks. The time-courses were decomposed into five frequency bands by discrete wavelet decomposition. The amplitude of the frequency-specific activity was compared between groups. Furthermore, the preprocessed resting state images were decomposed by wavelet analysis into five specific frequency bands voxel-wise. The voxel-wise amplitudes were compared between groups by non-parametric permutation test. RESULTS: In the MWA-Control comparison the discrete wavelet decomposition found alterations in the lateral visual network. Higher activity was measured in the MWA group in the highest frequency band (0.16-0.08 Hz). In case of the MWA-MWoA comparison all networks showed higher activity in the 0.08-0.04 Hz frequency range in MWA, and the lateral visual network in in higher frequencies. In MWoA-Control comparison only the default mode network revealed decreased activity in MWoA group in the 0.08-0.04 Hz band. The voxel-wise frequency specific analysis of the amplitudes found higher amplitudes in MWA as compared to MWoA in the in fronto-parietal regions, anterior cingulate cortex and cerebellum. DISCUSSION: The amplitude of the resting state fMRI activity fluctuation is higher in MWA than in MWoA. These results are in concordance with former studies, which found cortical hyperexcitability in MWA.

Evidence for Plastic Processes in Migraine with Aura: A Diffusion Weighted MRI Study.

Background: Formerly white matter abnormalities in a mixed group of migraine patients with and without aura were shown. Here, we aimed to explore white matter alterations in a homogeneous group of migraineurs with aura and to delineate possible relationships between white matter changes and clinical variables. Methods: Eighteen patients with aura, 25 migraine patients without aura and 28 controls were scanned on a 1.5T MRI scanner. Diffusivity parameters of the white matter were estimated and compared between patients' groups and controls using whole-brain tract-based spatial statistics. Results: Decreased radial diffusivity (p < 0.036) was found bilaterally in the parieto-occipital white matter, the corpus callosum, and the cingular white matter of migraine with aura (MwA) patients compared to controls. Migraine without aura (MwoA) patients showed no alteration compared to controls. MwA compared to MwoA showed increased fractional anisotropy (p < 0.048) in the left parieto-occipital white matter. In MwA a negative correlation was found between axial diffusivity and disease duration in the left superior longitudinal fascicle (left parieto-occipital region) and in the left corticospinal tract (p < 0.036) and with the number of the attacks in the right superior longitudinal fascicle (p < 0.048). Conclusion: We showed for the first time that there are white matter microstructural differences between these two subgroups of migraine and hence it is important to handle the two groups separately in further researches. We propose that degenerative and maladaptive plastic changes coexist in the disease and the diffusion profile is a result of these processes.

Kynurenic Acid Inhibits the Electrical Stimulation Induced Elevated Pituitary Adenylate Cyclase-Activating Polypeptide Expression in the TNC.

BACKGROUND: Migraine is a primary headache of imprecisely known mechanism, but activation of the trigeminovascular system (TS) appears to be essential during the attack. Intensive research has recently focused on pituitary adenylate cyclase-activating polypeptide (PACAP) and the kynurenine systems as potential pathogenic factors. AIM: We investigated the link between these important mediators and the effects of kynurenic acid (KYNA) and its synthetic analog (KYNA-a) on PACAP expression in the rat trigeminal nucleus caudalis (TNC) in a TS stimulation model related to migraine mechanisms. METHODS: Adult male Sprague-Dawley rats were pretreated with KYNA, KYNA-a, the NMDA receptor antagonist MK-801, or saline (vehicle). Next, the trigeminal ganglion (TRG) was electrically stimulated, the animals were transcardially perfused following 180 min, and the TNC was removed. In the TNC samples, 38 amino acid form of PACAP (PACAP1-38)-like radioimmunoactivity was measured by radioimmunoassay, the relative optical density of preproPACAP was assessed by Western blot analysis, and PACAP1-38 mRNA was detected by real-time PCR. RESULTS AND CONCLUSION: Electrical TRG stimulation resulted in significant increases of PACAP1-38-LI, preproPACAP, and PACAP1-38 mRNA in the TNC. These increases were prevented by the pretreatments with KYNA, KYNA-a, and MK-801. This is the first study to provide evidence for a direct link between PACAP and the kynurenine system during TS activation.

Ipsilateral Alteration of Resting State Activity Suggests That Cortical Dysfunction Contributes to the Pathogenesis of Cluster Headache.

The pathomechanism of cluster headache (CH) is not entirely understood, but central and peripheral components were suggested. A recent report showed that transcranial magnetic stimulation measured cortical excitability was increased in the hemisphere ipsilalteral to the pain. In the current study we set out to investigate the amplitude of resting brain fMRI activity to find signatures of the increased excitability. High resolution T1 weighted and resting state functional MRI images were acquired from seventeen patients with CH in pain free period and from twenty-six healthy volunteers. Patients' data were normalized (e.g. inverted along the midsagittal axis) according to the headache side. Independent component analysis and a modified dual regression approach were used to reveal the differences between the resting state networks. Furthermore, the timecourses were decomposed into five frequency bands by discrete wavelet decomposition and were also re-regressed to the original data to reveal frequency specific resting activity maps. Two of the identified resting state networks showed alterations in CH. When the data were inverted to have patients' headaches on the left, the ipsilateral attention network showed increased connectivity in 0.08-0.04 Hz frequency band in the in CH group. In the same dataset, cerebellar network showed higher functional connectivity in 0.02-0.01 Hz range in the ipsilateral cerebellum. When the data of patients having headache on the left were inverted to the right, similar increased signal was found in the ipsilateral attention network in 0.08-0.04 Hz band. The cerebellar network showed increased connectivity in the cerebellum in 0.02-0.01 Hz band in patients. The Fourier analysis of these area revealed increased power in CH at all cases. Our results showed alterations of brain functional networks in CH. The alterations of resting state activity were found in the hemisphere ipsilateral to the pain, signifying the altered cortical processing in the pathomechanism of CH.

Release of PACAP-38 in episodic cluster headache patients - an exploratory study.

BACKGROUND: Activation of the trigeminal-autonomic reflex, involving the trigeminal ganglion, the superior salivatory nucleus and the sphenopalatine ganglion (SPG) is crucial in the pathophysiology of cluster headache (CH). Since pituitary adenylate cyclase-activating polypeptide-38 (PACAP-38) is present both in the SPG and the trigeminal ganglion (TG) and its role in migraine has been described, our aim was to determine the plasma PACAP-38 levels in different phases of episodic CH (ECH). Peripheral cubital fossa blood samples were taken during the ictal and inter-bout periods of male ECH patients and from age-matched healthy controls (n = 9). Plasma PACAP-38-like immunoreactivity (LI) was measured with specific and sensitive radioimmunoassay. FINDINGS: Significantly lower plasma PACAP-38-LI was detected in the inter-bout period of ECH patients than in healthy controls. However, PACAP-38 was significantly elevated in the plasma during CH attacks as compared to the inter-bout phase in the same subjects (n = 5). CONCLUSIONS: This exploratory study suggests that PACAP-38 may be released during the attacks of ECH. Further patients and long-term follow-up are necessary to reveal its function.

Utility of different outcome measures for the nitroglycerin model of migraine in mice.

INTRODUCTION: Majority of the work for establishing nitroglycerin (NTG)-induced migraine models in animals was done in rats, though recently some studies in mice were also reported. Different special formulations of NTG were investigated in various studies; however, NTG treated groups were often compared to simple saline treated control groups. The aim of the present studies was to critically assess the utility of a panel of potential outcome measures in mice by revisiting previous findings and investigating endpoints that have not been tested in mice yet. METHODS: We investigated two NTG formulations, Nitrolingual and Nitro Pohl, at an intraperitoneal dose of 10mg/kg, in comparison with relevant vehicle controls, and evaluated the following outcome measures: light aversive behaviour, cranial blood perfusion by laser Doppler imaging, number of c-Fos- and neuronal nitrogen monoxide synthase (nNOS)-immunoreactive neurons in the trigeminal nucleus caudalis (TNC) and trigeminal ganglia, thermal hyperalgesia and tactile allodynia of the hind paw and orofacial pain hypersensitivity. RESULTS: We could not confirm previous reports of significant NTG-induced changes in light aversion and cranial blood perfusion of mice but we observed considerable effects elicited by the vehicle of Nitrolingual. In contrast, the vehicle of Nitro Pohl was apparently inert. Increased c-Fos expression in the TNC, thermal hyperalgesia, tactile allodynia and orofacial hypersensitivity were apparently good endpoints in mice that were increased by NTG-administration. The NTG-induced increase in c-Fos expression was prevented by topiramate but not by sumatriptan treatment. However, the NTG-induced orofacial hypersensitivity was dose dependently attenuated by sumatriptan. DISCUSSION: Our results pointed to utilisable NTG formulations and outcome measures for NTG-induced migraine models in mice. Pending further cross-validation with positive and negative control drugs in these mouse models and in the human NTG models of migraine, these tests might be valuable translational research tools for development of new anti-migraine drugs.