The Changes And Significances Of NRG1-ErbB4Signal In Brain Tissues Of Fmr1Knockout Mice

The fragile X syndrome is the most frequent form of inherited mental retardation inhumans, with an incidence of1in4000males and1in8000females.The molecular basisfor this disease is the unstable amplified of a triplet repeat （CGG）,in the5’ untranslatedregion and abnormal methylation of cytosine nucleotides of fragile X mental retardationgene （FMR1）, resulting in gene transcriptional silence and gene product fragile X mentalretardation protein （FMRP） reduced or missing.The main clinical manifestations includevarying degrees of mental retardation, attention deficit hyperactivity and autism,20%-25%of patients suffered from epilepsy.Recent studies has shown that thehypofunction of the inhibitory interneurons leads to enhanced excitability in cerebralcortex have linked to the pathogenesis in FXS.We also found the reduced number of GADand PV positive neurons, as well as decreased current density of type I voltage-gatedsodium channel （Nav1.1） on interneuron in Fmr1knockout mice.Our research group alsofound the area of PV cells reduce in cortex, temporal cortex, piriform cortex compared tothe age-match WT mice in2weeks Fmr1KO mice. There is significant increased in CA2,CA3, striate cortex,temporal auditory cortex and piriform cortex in KO^4wmice comparedwith WT^4w.And Our research group also found GABAAR α5protein expression and inmRNA level were lower than that of the WT mice in Fmr1KO mice at the age of14-17days.Studies have shown that the NRG1（Neuregulin1） and its receptor ErbB4play a vital role in neuronal development、synaptic function and plasticity、 neurotransmitter and theregulation of N-methyl-D-aspartate, GABAA、acetylcholine receptor subunit expressionas well as maintain the balance of the neuron circuits.Therefore we speculate the changeof NRG1-ErbB4signal cause PV positive neurons abnormalities in the number andstructural in Fmr1KO mouse and increase seizures susceptibility in Fmr1KO mouse; Ourresearch study the expression of NRG1、ErbB4neurons and ErbB4distribution on thePV-positive interneurons to explore the role of NRG1-ErbB4signal in epilepsy increasedsusceptibility in the FXS.Materials and MethodsFVB strains of mice reared under natural light conditions of19²1℃, free accessto food and water.We extract DNA from tail for PCR to dentify the genotypes.Then wedetect the NRG1and ErbB4number of neurons by immunohistochemical method （eachgroup sacrificed6mice） and protein by Western blot （each group sacrificed3mice）; Wedetect ErbB4protein express on the PV-positive neurons through doubleimmunofluorescence labeling technique（each group sacrificed3mice）. All mouse wererandomly selected. male Fmr1knockout mice after the birth of two weeks and four weeksweeks （the knockout mouse, KO）(KO^2wand KO^4w)as experimental groups and the sameage, male wild-type (wild type, WT, WT^2wand WT^4w) mice served as controls.Results1.Comparison of the number of NRG1-positive cells in WT and KO mice braintissuesThe numbers of NRG1-positive cells in the regions of CA1and CA3of thehippocampus and the cortex of Fmr1KO mice at the age of2and4weeks weresignificantly less than those in the age-matched WT mice（P﹤0.05）.Conversely, thenumbers of NRG1-positive cells was increased in the dentate gyrus of Fmr1KO mice（P﹤0.05）.2.Comparison of the number of ErbB4-positive cells in WT and KO mice braintissuesThe numbers of ErbB4-positive cells in the regions of CA1、CA3and dentate gyrus of the hippocampus and the cortex of Fmr1KO mice at the age of2weeks weresignificantly less than those in the age-matched WT mice（P﹤0.05）,The numbers ofErbB4-positive cells in the regions of CA3of the hippocampus and the cortex of Fmr1KO mice at the age of4weeks were significantly less than those in the age-matched WTmice（P﹤0.05）.No significant different was found in hippocampus CA1and dentate gyrusbetween WT^4wand KO^4w.3. Comparison of the level of NRG1and ErbB4protein in WT and KO mice braintissuesThe level of NRG1and ErbB4protein in the hippocampus and cerebral cortex inFmr1KO mice at the age of2and4weeks were decreased when compared with theage-match WT mice（P﹤0.05）.4. Comparison of the number of ErbB4protein on the parvalbumin-positive cells inWT and KO mice brain tissuesErbB4expression on PV-positive neurons were reduced in cortex and CA1、CA3ofthe hippocampus of KO^2wand KO^4wmouse brain compared with WT^2wand WT^4wrespectively. No significant different was found in hippocampus dentate gyrus betweentwo groups.ConclusionThe numbers and amount level of NRG1、ErbB4positive neurons and ErbB4expression on PV-positive neurons were reduced in cortex and hippocampus of Fmr1KOmouse brain, We prompt the abnormal NRG1-ErbB4signal related to reduction of the PV-positive GABAergic interneuron.,and it may play an important role in the increasedepilepsy susceptibility in FXS...