| Objective：With the development of technology and economy, thetrend of aging increasingly obvious. Aging and age-associated dementianot only impose heavy burdens on the individuals and families, but alsoimpede the development of social and economy. Thus the study ofimprove cognitive function, delay brain aging and prevent age-associateddisease have been considered an important mission in this aging society.Repetitive transcranial magnetic stimulation(rTMS) is a noninvasive brainstimulation with minimal side effects, thus it has become increasinglyimportant in brain research and is widely used in neurologic andpsychiatric disorders. The exposure of human brain to a specific pattern ofrTMS could modulates brain function and induced persistent neuronalexcitability in the hippocampal, and lead to lasting effects that are thoughtto be due to changes in synaptic plasticity such as long term depression(LTD) and long term potentiation (LTP). Apparently, the expression ofseveral synaptic plasticity associated genes such as immediate-earlygenes(IEGs) are strongly induced during LTP/LTD-inducing processes.Yet the molecular and cellular basis underlying synaptic plasticity and theregulatory effects on aging brain remains dimly illuminated. In a series ofexperiments we aimed to characterize the effects of different pattern ofrTMS on neuronal activity and synaptic plasticity associated geneexpression through Morris water maze and PCR array analysis and toevaluate the potential correlations of these modifications with learningand memory in aging brain, thus provide fundamental basis for thetreatment of neurodegenerative disease.Methods:(1) Animals:40aged Kunming mice(15-months-old).(2) Methods:①rTMS：Forty aged Kunming mice were divided into threegroups depending on their cognitive function: sham,5Hz20%maxexport intensity and30%max export intensity. Two sessions of rTMSconsist of1000pulses in10trains were delivered on14consecutive days.Sham group were treated by the reverse side of the coil.②Morris watermaze: It was done on the first day after rTMS, the escape latency wasrecorded on five continues days, and the number of times that each mousecrossed the previous platform location was recorded as a measure ofplatform location retention on the sixth day.③Hippocampal RNAExtraction: In the seventh day of rTMS, the mice were sacrificed and thehippocampus were quickly obtained and stored at–80°C. Total RNAwas extracted from hippocampal tissues using RNeasy Mini Kitaccording to the manufacturer’s protocol.④Hippocampal RNA reversetranscription: RNA was reversed to single strand cDNA using RT2FirstStrand Kit with1μ g RNA each group.⑤PCR array analysis：Differential expression of synaptic plasticity associated genes after rTMSwas analyzed with the Mouse Synaptic Plasticity RT2Profiler PCR Arraywhich contains84synaptic plasticity associated genes,5housekeepinggenes and7quality control genes. The results were analysed withSABiosciences PCR Array data analysis Excel templete.⑥Morris watermaze was determined by ANOVA for repeated measurement and one-wayANOVA using SPSS13.0software. All data were presented as mean±SD. A P value cut-off0.05was considered significant.Results: Effects of rTMS on learning and memory of aged Kunmingmice: The results of Morris water maze showed that the mean escapelatency of both the rTMS treatment groups were significantly shortercompared with sham group(P<0.05), which indicated that learning andspatial memory of aged Kunming mice were improved after rTMSadministration. More over,5Hz30%intensity group(P<0.01) was moresignificant than5Hz20%intensity group(P<0.05). The number ofcrossing original platform of both rTMS groups were significantly increased(P<0.05). As the same as the mean latency results,5Hz30%intensity group(P<0.01) was more significant than5Hz20%intensitygroup(P<0.05).(2)Effects of rTMS on synaptic plasticity associatedgenes by the use of PCR array: The84genes were classified into tengroups associated with synaptic plasticity, which consist ofImmediate-Early Response Genes (IEGs), LTP, LTD, cell adhesion,CREB cofactors, Extracellular matrix&proteolytic processing(EMPP),neuronal receptors(NR), postsynaptic density(PD), late response genesand other genes important for synapse remodeling. On the basis ofgenetic changes, we choose up regulation for1.5fold and downregulation for2fold as meaningful genes. In the5Hz30%intensity group,there are10genes were found to be up-regulated by at least1.5-fold,13genes down-regulated by at least2-fold Compared to the sham group.Whereas in the5Hz30%intensity group, there are9genes were found tobe up-regulated by at least1.5-fold,25genes down-regulated by at least2-fold. The maximum up or down regulated gene are all in5Hz30%intensity group. Among these regulated genes, Junb、Grip1in5Hz30%intensity group up-regulated11.41-fold,2.73-fold respectively， Inhbadown-regulated for3.45-fold. While Junb, Nr4a1、Grip1、Camk2g in5Hz30%intensity group up-regulated for3.73-fold、3.47-fold、1.74-fold、1.76-fold respectively，and Inhba down-regulated for2.27-fold.Conclusion:(1) High frequency rTMS can significantly improvespacial memory of aged Kun ming mice, which indicated that rTMS iseffective in improving cognitive function of brain aging.(2) Highfrequency rTMS lead to the alterant of synaptic plasticity associatedgenes, and the improvement in cognitive function by differentintensities of stimulation may involve different gene regulation althoughthey share most of the similarities. |
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