MERTK in the rat trigeminal system: a potential novel target for cluster headache?

The trigeminal system is key to the pathophysiology of migraine and cluster headache, two primary headache disorders that share many features. Recently, MER proto-oncogene tyrosine kinase (MERTK), a cell surface receptor, was strongly associated with cluster headache through genetic studies. Further, the MERTK ligand galectin-3 has been found to be elevated in serum of migraine patients. In this study, MERTK and MERTK ligands were investigated in key tissue to better understand their potential implication in the pathophysiology of primary headache disorders. Immunohistochemistry was used to map MERTK and galectin-3 expression in rat trigeminal ganglia. RT-qPCR was used to assess MERTK gene expression in blood, and ELISA immunoassays were used for MERTK ligand quantification in serum from study participants with and without cluster headache. MERTK gene expression was elevated in blood samples from study participants with cluster headache compared to controls. In addition, MERTK ligand galectin-3 was found at increased concentration in the serum of study participants with cluster headache, whereas the levels of MERTK ligands growth arrest specific 6 and protein S unaffected. MERTK and galectin-3 were both expressed in rat trigeminal ganglia. Galectin-3 was primarily localized in smaller neurons and to a lesser extent in C-fibres, while MERTK was found in satellite glia cells and in the outer membrane of Schwann cells. Interestingly, a strong MERTK signal was found specifically in the region proximal to the nodes of Ranvier. The overexpression of MERTK and galectin-3 in tissue from study participants with cluster headache, as well as the presence of MERTK in rat peripheral satellite glia cells and Schwann cells in the trigeminal ganglia, further highlights MERTK signalling as an interesting potential future therapeutic target in primary headache.

The genetics and chronobiology of cluster headache.

BACKGROUND/HYPOTHESIS: Cluster headache displays uniquely rhythmic patterns in its attack manifestation. This strong chronobiological influence suggests that part of the pathophysiology of cluster headache is distinctly different from migraine and has prompted genetic investigations probing these systems. METHODS: This is a narrative overview of the cluster headache chronobiological phenotype from the point of view of genetics covering existing knowledge, highlighting the specific challenges in cluster headache and suggesting novel research approaches to overcome these. RESULTS: The chronobiological features of cluster headache are a hallmark of the disorder and while discrepancies between study results do exist, the main findings are highly reproducible across populations and time. Particular findings in subgroups indicate that the heritability of the disorder is linked to chronobiological systems. Meanwhile, genetic markers of circadian rhythm genes have been implicated in cluster headache, but with conflicting results. However, in two recently published genome wide association studies two of the identified four loci include genes with an involvement in circadian rhythm, MER proto-oncogene, tyrosine kinase and four and a half LIM domains 5. These findings strengthen the involvement of circadian rhythm in cluster headache pathophysiology. CONCLUSION/INTERPRETATION: Studying chronobiology and genetics in cluster headache presents challenges unique to the disorder. Researchers are overcoming these challenges by pooling various data from different cohorts and performing meta-analyses providing novel insights into a classically enigmatic disorder. Further progress can likely be made by combining deep pheno- and genotyping.

Cluster headache - clinical pattern and a new severity scale in a Swedish cohort.

Background The aim of this study was to investigate clinical features of a cluster headache cohort in Sweden and to construct and test a new scale for grading severity. Methods Subjects were identified by screening medical records for the ICD 10 code G44.0, that is, cluster headache. Five hundred participating research subjects filled in a questionnaire including personal, demographic and medical aspects. We constructed a novel scale for grading cluster headache in this cohort: The Cluster Headache Severity Scale, which included number of attacks per day, attack and period duration. The lowest total score was three and the highest 12, and we used the Cluster Headache Severity Scale to grade subjects suffering from cluster headache. We further implemented the scale by defining a cluster headache maximum severity subgroup with a high Cluster Headache Severity Scale score ≥ 9. Results A majority (66.7%) of the patients reported that attacks appear at certain time intervals. In addition, cluster headache patients who were current tobacco users or had a history of tobacco consumption had a later age of disease onset (31.7 years) compared to non-tobacco users (28.5 years). The Cluster Headache Severity Scale score was higher in the patient group reporting sporadic or no alcohol intake than in the groups reporting an alcohol consumption of three to four standard units per week or more. Maximum severity cluster headache patients were characterised by higher age at disease onset, greater use of prophylactic medication, reduced hours of sleep, and lower alcohol consumption compared to the non-cluster headache maximum severity group. Conclusion There was a wide variation of severity grade among cluster headache patients, with a very marked impact on daily living for the most profoundly affected.

Screening of genetic variants in ADCYAP1R1, MME and 14q21 in a Swedish cluster headache cohort.

BACKGROUND: We have genotyped a Swedish cluster headache case-control population for three genetic variants representing the most significant markers identified in a recently published genome wide association study on cluster headache. The genetic variants were two common polymorphisms; rs12668955 in ADCYAP1R1 (adenylate cyclase activating polypeptide 1 receptor type 1), rs1006417, an intergenic variant on chromosome 14q21 and one rare mutation, rs147564881, in MME (membrane metalloendopeptidase). RESULTS: We screened 542 cluster headache patients and 581 controls using TaqMan real-time PCR on a 7500 fast cycler, and pyrosequencing on a PSQ 96 System. Statistical analysis for genotype and allele association showed that neither of the two common variants, rs12668955 and rs1006417 were associated with cluster headache. The MME mutation was investigated with pyrosequencing in patients, of whom all were wild type. CONCLUSION: In conclusion rs12668955 and rs1006417 do not impact the risk of developing cluster headache in the Swedish population. Also, rs147564881 does not seem to be enriched within the Swedish cluster headache patient group.

ReMo-SNPs: a new software tool for identification of polymorphisms in regions and motifs genome-wide.

Studies of complex genetic diseases have revealed many risk factors of small effect, but the combined amount of heritability explained is still low. Genome-wide association studies are often underpowered to identify true effects because of the very large number of parallel tests. There is, therefore, a great need to generate data sets that are enriched for those markers that have an increased a priori chance of being functional, such as markers in genomic regions involved in gene regulation. ReMo-SNPs is a computational program developed to aid researchers in the process of selecting functional SNPs for association analyses in user-specified regions and/or motifs genome-wide. The useful feature of automatic selection of genotyped markers in the user-provided material makes the output data ready to be used in a following association study. In this article we describe the program and its functions. We also validate the program by including an example study on three different transcription factors and results from an association study on two psychiatric phenotypes. The flexibility of the ReMo-SNPs program enables the user to study any region or sequence of interest, without limitation to transcription factor binding regions and motifs. The program is freely available at: http://www.neuro.ki.se/ReMo-SNPs/.

Genetic studies of the protein kinase AKT1 in Parkinson's disease.

The protein kinase AKT1 belongs to the Akt family and is a potent mediator of cell growth and survival and fully activated when phosphorylated. The AKT family has been found to be phosphorylated to a lesser extent in the dopaminergic cells of Parkinson's disease patients compared to control individuals, which might influence cell survival. Several publications support the implication of AKT1 in disorders of the dopaminergic system including bipolar disease and schizophrenia. In 2008 an association study performed in a Greek Parkinson's disease case-control material reported the identification of a protective AKT1 haplotype. Based on their work we have performed a replication study in a Swedish Parkinson's disease cohort. We genotyped the four single nucleotide polymorphims (SNPs): rs2494743, rs2498788, rs2494746 and rs1130214 in a case-control material consisting of 243 Parkinson patients and 315 controls. We did not find any associations with Parkinson's disease for either the individual SNPs or any of the haplotypes. In contrast to previously published results, our data do not support the hypothesis of genetic variants in AKT1 confering protection against Parkinson's disease.

Expression of multi-drug resistance 1 mRNA in human and rodent tissues: reduced levels in Parkinson patients.

The membrane transporter multi-drug resistance 1 (MDR1, P-gp) regulates the bioavailability of endogenous and exogenous compounds and has been implicated in disorders such as Parkinson's disease, cancer, epilepsy, human immunodeficiency virus disease, and inflammatory bowel disease. To promote further understanding of the role of MDR1 in disease, we have characterized cellular MDR1 mRNA expression in post-mortem human and fresh-frozen Sprague-Dawley rat tissues by using radioactive oligonucleotide probe in situ hybridization. We report MDR1 mRNA in human and rat endothelial cells of small vessels in the brain and pia mater. Mdr1 mRNA is also expressed in the blood vessel walls of rat sensory dorsal root and sympathetic ganglia. In peripheral tissues, we have observed MDR1 mRNA in human and rat liver and renal tubules and in human adrenal cortex and the epithelial lining of rat intestine. In female and male reproductive tissues of rat, strong gene activity has been found in steroid-hormone-synthesizing cells. Quantification of MDR1 mRNA in human striatum has revealed reduced levels in Parkinson patients compared with control individuals. The high expression of MDR1 mRNA in blood vessels of the nervous system, in tissues involved in absorption and excretion, and in tissues forming barriers to the environment support the physiological role of MDR1 as a regulator of intracellular levels of endogenous and exogenous compounds.

DJ-1 and UCH-L1 gene activity patterns in the brains of controls, Parkinson and schizophrenia patients and in rodents.

DJ-1 (PARK7) has been implicated in early onset and familial cases of Parkinson's disease (PD). We therefore mapped cellular activity patterns of the DJ-1 gene in human and rodent brain tissue with radioactive in-situ hybridization. In all three mammals mRNA expression was restricted mainly to neurons in all regions analyzed. White matter, such as crus cerebri and capsula interna appeared negative, suggesting that glial cells express DJ-1 at levels below the detection limit of our method. We compared DJ-1 mRNA expression to the neuronal marker UCH-L1, which has also been implicated in PD, and found lower levels for DJ-1 but very similar patterns of expression. Measurement of the signal intensity revealed that human frontal cortex of control cases expressed DJ-1 mRNA more abundantly than other regions such as substantia nigra in the midbrain. Comparing DJ-1 expression in dopamine neurons on hemi-sections from controls and patients we could not detect any difference between 14 controls, 8 idiopathic Parkinson and 5 schizophrenia cases. Of note, DJ-1 is expressed in several other tissues such as the liver, gastrointestinal tract, adrenal and pituitary gland and during embryonic development, while UCH-L1 has a strictly neuronal expression also outside the CNS. We conclude that DJ-1 and UCH-L1, like other genes linked to PD, are not expressed specifically in DA neurons, but instead generally in neurons. The abundant expression of DJ-1 in certain peripheral tissues and of UCH-L1 in peripheral neurons may also be of relevance for the spectrum of symptoms in different forms of PD.