Change Of Voltage-gated Sodium Channel α Subunit MRNA In Brain Of Spontaneously Epileptic Rat

The relationship between the change of voltage - gated sodium channel a subunit and pathogenesis of epilepsy is very closely. This report used genetic epileptic animal model, spontaneously epileptic rat ( SER) , as experimental animal , and compared voltage - gated I , II and III sodium channel a subunit mRNA relative amount of SER with that of normal Wistar rats in brain. To find the exact relationship between genetic epilepsy pathogenesis and encoding voltage - gated sodium channel sequence, and provide theoretical foundation for gene therapy of epilepsy.MethodsFive SER and Wistar rats were in each group. They were killed by decapitation , and the brain were removed rapidly, frozen in liquid vapors and spliced into 400 um coronal section. Under binocular microscopic needles punches were sampled from four different regions: cortex, dentate gyrus, hippocampus CA1, and CA3. Total RNA extracted from these four regions tissue, with RNA PCR kit performed PCR protocol 1, 2 and 3, respectively. Amplified products according to PCR 1 protocol were run agarose gel electrophoresis, with ultraviolet scanning analysis compared voltage - gated sodium channel a subunit mRNA relative amount of SER with that of Wistar rats in four different regions of brain. Amplified products according to PCR 2 and/or 3 were performed specific restriction enzyme analysis, Msp I for NaCh I , BamH I for NaCh II A, Sfu I for NaCh II N, Dra I for NaCh III, Nde I for NaCh III A, and Sac I for NaCh III N, respectively. Digested products were run agarose gel electrophoresis and with ul-traviolet scanning detected the change of subtype I , IIA, II N, III A, and III N.ResultsSER all types of sodium channel expressed a little higher than control groups in cortex, dentate gyrus and CA1, but had no significant difference ( P> 0.05, P value is 0.36, 0. 24, and 0.41, respectively ) , and were the same with control rats in CA3. Digested by specific restriction enzyme, SER type I ,II A, II N, and III sodium channel percentage constitution in the whole sodium channel are all similar to Wistar rats. Specific amplified products for type III sodium channel under restriction analysis, showed that III A and III N fragments were present in SER brain simultaneously. But during brain development, subtype III N expressed highly in embryonic stage, and disappeared postnatal 10 days, subtype III A replaced it gradually. So III N should not have appeared in a-dult Wistar rats. These indicated that SER subtype IIIN mRNA increased significantly (P < 0.01).ConclusionsIn the amplified voltage - gated sodium channel a subunit coding sequences , there was no large deletion or mutation in SER brain. But mRNA mutually exclusive splicing of type III sodium channel two exons, III A and III N, in SER brain was not regulated correctly during brain development. This might result in discharging excessively by neuron, but it required the evidence verified by voltage - gated sodium channel electrophysiological studies.