| Objective:1.To observe the gene expression profiling in trigeminal nucleus caudalis of the alert rats after electrical stimulation of dura matter adjacent to superior sagittal sinus(SSS),and to find the differential expressed genes.2.To validate some changed genes that are related to pathophysiology of migraine by real-time quantified PCR and to determine the possible molecular mechanism of migraine.3.To further validate the gene expression of AKAP5 by the western-blot technique.Methods:1.The gene expression profiling of the electrical stimulation model in trigeminal nucleus caudalis was studied using Affymetrix Rat Genome 230 2.0 Oligo microarrays.Rats were rapidly decapitated and the medulla and high-level cervical cord(to C2) were removed from the skull after electrical stimulation. Then RNA samples were extracted.After hybridization of the cRNA to the Affymetrix Rat Genome 230 2.0 gene chip,the chip was washed,stained and scanned on a GeneChip Scanner 3000.GeneChip operating software(GCOS) was used to analyze the presentation.Then SAM software was used to find the differential expressed genes between each group.Functional classification for differential genes was further performed by Molecule Annotation System.2.Gene expression validation for 10 genes was performed by Real-Time quantified polymerase chain reaction(PCR).Functional analysis of differential expressed genes were performed by bioinformatics analysis.Molecular mechanism of pain involved in migraine attack was evaluated.3.27 rats were randomly divided into four groups including control group(n=4), groups of electrical stimulation for 30 minutes(n=6),60 minutes(n=6),120 minutes(n=5) and group of morphine plus electrical stimulation for 120 minutes(n=6).Rats were rapidly decapitated,and the medulla and high-level cervical cord(to C2) were removed from the skull after electrical stimulation..AKAP5 was detected in the medulla and high-level cervical cord by western blot technique.Results:1.According to the "Present" by detection p-value＜0.05,21052.33±194.50 transcripts were detected among the 31099 genomic probe sets in control group, in proportion to 67.69%for total transcripts,meanwhile 20812.67±130.89 transcripts were detected in model group and had a proportion of 66.92%. Then the differential expressed genes were screened between two groups using GCOS,SAM and MAS system.Compared with control group,there were 28 down-regulated genes and 16 up-regulated genes in model group,which were involved in retinol metabolism,adherens junction,cell adhesion molecules (CAMs),porter activity,electrochemical potential-driven transporter activity, cytoskeletal protein binding,actin binding,enzyme inhibitor activity, mitochondrion,Krebs-TCA_Cycle,coenzyme metabolism,organelle membrane,protein folding,binding,intracellular protein transport,kinase activity,G_protein_signaling,inorganic anion transport,monovalent inorganic cation transport,integral to plasma membrane,intrinsic to plasma membrane, adipogenesis,metabolism,tryptophane_metabolism,oxidoreductase activity, extracellular matrix(sensu Metazoa),extracellular matrix,extracellular region, development,signal transducer activity,lipid binding,cytoplasm,nervous system development,cell adhesion,system development,cell_cycle, striated_muscle_contraction,anion transport,ion transport,transporter activity, metal ion transport,neurotransmitter:sodium symporter activity,symporter activity,neurotransmitter transport pathways.2.Of the 10 genes analysed by RT-PCR,the direction and degree of change matched almost perfectly to the microarray data.3.AKAP5 was expressed in the bulb and high cervical cord.There were no significant deviation of AKAP5 expression among the five groups.Conclusions: 1.Electric stimulation of dura matter adjacent to superior sagittal sinus(SSS) caused 44 genes expression changes of trigeminal nucleus caudalis,among which were several related with porter activity and electrochemical potential-driven transporter activity,3 genes were involed in each pathway. Most of the genes were involed in more than 2 pathways.They may contribute to the attack of migraine.2.RT-PCR confirmed the microarray results for several of the transcripts.3.There was no significant deviation for AKAP5 expression between control and electrical stimulation groups.Morphine has no effect on AKAP5 gene expression.
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