Roles Of NRG1/ErbB4 Pathway In Parvalbumin Positive Interneurons In Cognitive Impairment In Mice With Cornelia De Lange Syndrome

Backgroud:Cornelia de Lange Syndrome(CdLS;MIM#122470,300590),also known as Amsterdam dwarfism,is a rare genetically heterogeneous disorders characterized by distinctive growth retardation,intellectual disability(IQ<40),and behavioral problems.In addition,these patients are incapacitated and thus bring much burden for their families and society.CdLS is hereditarily congenital malformation caused by the heterozygous mutation of cohesin-encoding genes including Nipped-B-like protein(NIPBL),regulator of cohesion maintenance(PDS5),structural maintenance of chromosomes 1(SMC1),structural maintenance of chromosomes 3(SMC3)and double-strand-break repair protein(RAD21).The cohesion complex plays an important role in the segregation of sister chromatid during cell division and regulation of gene expression and DNA repair.Therefore,the mutation of cohesion complex seriously impairs the early embryonic development.Approximately 50%of patients with CdLS are accompanied with NIPBL mutation.Nevertheless,little is known about the effects of NIPBL mutation in learning and memory in CdLS as well as its underlying mechanism.Hippocampus is very important in learning and memory,and its synaptic plasticity is involved in memory induction and formation.Upon exogenous stimulation,the hippocampus synapse undergoes the modulation to receive new information.Among the various types of synaptic plasticities,long-time potentiation(LTP)is a classic model of learning and memory,while long-term depression(LTD)involves in the consolidation and retrieve of memory.NRG1/ErbB signaling pathway is not only required for the development of central nervous system and brain function,but also for maintaining high level of GABAergic system including the interneuron.NRGl/ErbB4 pathway is critical for the synapses formation,maturation and transmission.Additionally,it is one of the important mechanisms involving in neurodevelopmental and mental disorders.CdLS is overlapping with these disorders.It has been demonstrated that NRGl/ErbB4 pathway in the GABAergic interneuron could inhibit the induce of LTP in the hippocampus.Therefore,we would like to reveal the role of dysregulation of NRG1/ErbB4 pathway in PV positive interneurons in CdLS.Up to now,it has not been reported whether NRGl/ErbB4 pathway was involved in the cognitive impairment in the CdLS.Method:We firstly cultured NIPBLflox/+,EIIA-Cre mice and NIPBLflox/+,PV-Cre mice based on the Cre-LoxP recombinant system.The weight of male mice was recorded.Mice genotypes were examined by PCR.Behavioral tests including Morris water maze and Shuttle box were conducted to evaluate learning and memory.Real time-PCR,immunohistochemistry and western blot were used to analyse the expression of NIPBL,NRG1,ErbB4,and PSD-95 in the cortex and hippocampus.Patch clamp technique was applied to measure the LTP and NRGl/ErbB4 pathway in the mice hippocampus.Result:NIPBLflox/+,E ? A-Cre mice displayed slow growth.NIPBLflox/+,E ?A-Cre mice and NIPBLflox/+,PV-Cre were generated as shown by PCR.In Morris water maze test,on the third training day the NIPBLflox/+PV-Cre mice(n=10)took more time(69.2 ± 34.6s vs 48.3±33.6s,P=0.167,t-test)and a longer swimming distance(1670.0±710.6cm vs 968,6±679.9cm,P=0.03,t-test)compared with the control group,while swimming velocity was almost the same(21.4±8.7cm/s vs 25.8±7.4 cm/s,p=0.167,t-test).On the fifth day,the munbers of cross platform(3.6 ± 1.8 vs 1.6 ± 1.2,P=0.01,t-test)were decreased.In addition,shuttle box experiments revealed that after 5 days training the escape latency in the NIPBLflox/+,PV-Cre mice(n=10)were longer(4.02 ±1.17s vs 2.8 ±0.84s,P=0.012,t-test)than in the control group.Compared with the control group,the mRNA expression of NIPBL was decreased;the mRNA and protein expression of NRG1,ErbB4 and PSD-95 were reduced significantly(P<0.05).We found that the down-regulated expression of NRG1,ErbB4,PSD-95 were partially recovered in the trained mice compared with those mice without training.Patch clamp assay showed that basal synaptic transmission was not affected,but LTP in NIPBLflox/+,PV-Cre mice was larger than that in the control group,which was partially reversed after NGR1 intervention(P<0.01,one way anova).These results indicated that dysregulation of NRG1/ErbB4 pathway in the hippocampus could probably lead to the cognitive impairment in CdLS mice.Conclusion:1.A mouse model of CdLS was generated using Cre-LoxP system.2.Compared with the control,in CdLS mice NRG1,ErbB4 and PSD-95 downregulation acompanied with a learning impairment and larger LTP in hippocampus.It implicated the dysregulation of NRG1/ErbB4 pathway in parvalbumin positive interneurons could be one of the pathogenesis of CdLS.3.NRG1 could partially reverse enhanced LTP in CdLS mice and be a potential therapeutic strategy.Significance:Our study would contribute to understanding the role of NIPBL in CdLS and CdLS-related hereditary diseases such as autism,epilepsy and autolesionism..In addition,our investigation delighted the role of NRG1/ErbB4 pathway in the GABAergic interneurons in CdLS,which might provide novel therapeutic strategy for such metal diseases in childhood.