Mechanism Of Vascular Headache Induced By Electrical Stimulation On Superior Sagittal Sinus Of Rats

Objective1,By means of electrical stimulation on superior sagittal sinus (SSS) of waking rats, combined with immunohistochemistry and immunofluorescence, the distribution of Fos-immunoreactive neurons in brain was systematically observed in order to find the active regions participating nociception during the stimulation, meaningwhile, the relationship between the Fos-immuoreactive neurons and the 5-HT-immunoreactive neurons was studied.2,To investigate the effect of flunarizine on ethology and Fos expression in the mode of electrical stimulation in SSS in waking rat.3,Protemic analysing of caudal subnucleus of the trigeminal nucleus caudalis of the waking rats after electrical stimulation on SSS was performed to search for the differential protein expression and explore possible mechanims of the vascular headache.Methods1,The male SD rats were randomly divided into experimental group and control group. After exposure of SSS, rats in the experimental group were received electrical stimulation on the dura mater. Immunohistochemistry was then utilized to label Fos expressing neurons and immunofluorescence was to label Fos or/and 5-HT expressing neurons, at the same time, the schematic map illustrating the distribution of Fos-immunoreactive neurons was drawn.2,After exposure of SSS of SD rats, self-control was employed to compare the behavior change after intraperitoneal injection of flunarizine with different concentration. Then the rats were randomly divided into experimental group (intraperitoneally flunarizine) and control group (intraperitoneally physiological saline) to immunochemically invesgate and compare the Fos expression between both groups and with the results from the first expriment.3,SD rats were randomly divided into experimental group and control group. After exposure of SSS, rats in the experimental group were received electrical stimulation on the dura mater near SSS. Then protemics study (protein electrophoresis and mass chromatographic analysis) was conducted for regions of trigeminal nucleus caudal part to search for differential expressed protein.Results1. After the electrical stimulation the Fos-immunoreactive neurons were observed in many areas in the brain, which were upper cervical spinal dorsal horn, spinal trigeminal nucleus caudalis, raphe nuclei, periaqueductal gray, interpeduncular nucleus and hypothalamus. Double immunofluorescence staining indicated that 5-HT-immunoreactive neurons existed mainly in the raphe nuclei and periaqueductal gray. Although Fos-immunoreactive neurons and 5-HT-immuroreactive neurons overlapped in the raphe nuclei and periaqueductal gray, only few Fos/5-HT co-localized neurons were found in the raphe magnus nucleus.2,The number of head shaking and grooming of rats with equal and high concentration of flunarizine was significantly reduced compared to low concentration and physiological saline.The distribution of Fos-immunoreactive neurons in the brain of rats in flunarizine injection group was the same with those in physiological saline injection group and normal group (experiment first), however, the Fos expression decreased in caudal subnucleus of trigeminal nuclei and increased in periadqueductal gray in rat brain of flunarizine injection group.3,After the two-dimensional gel electrophoresis, there were 1562 pairs of protein plot matched, among which 735 plot increased and 827 decreased.9 of the protein plot were selected for UPLC-ESI-MS/MS and they were GTP-binding nuclear protein Ran, lysophospholipase 2, glutathione S-transferase P, succinate semialdehyde dehydrogenase, Haloacid dehalogenase-like hydrolase domain-containing protein 2, transgelin, ATPase, H transporting, lysosomal V1 subunit G1, Septl1 protein and rCG55630, isoform CRA_a.Conclusions1. Several areas in the brain are involved in the generation and development of vascular headache in the mode of electrical stimulation on superior sagittal sinus. Except for nuclei modulating and transmitting nociceptive information, those relating to both motion and autonomic nerve regulation take part in the above process. In process of generation and development of vascular headache in the mode of electrical stimulation on SSS, the chemical property of the activated neurons is still need to further investigate. Some 5-HT immunoreactive neurons are activated, which means their participating in the process and may partially relate to the activation of descending antinociceptive system.2,The waking rat mode of electrical stimulating on SSS responed well to flunarizine for ethological parameters changed after injection of flunarizine indicating that pain was relieved, which further testified the feasibility of the mode. At the same time, the caudal subnucleus of trigeminal nuclei and periadequeductal gray were presumed to take part in the process of pain modulation from flunarizine for the Fos expression changed after injection of flunarizine, but specific mechanism was still unclear and need to explore.3,After the electrical stimulation on SSS, protemics of trigeminal nucleus caudalis were changed, indicating increased caryoplasm-cytoplasm transportion, increased anti-apoptosis and induced S-glutathionylation and decreased GABA degradation and so on, which may participate in the generation and development of the vascular headache.