Objective To explore the effect of electroacupuncture(EA)on learning-memory ability and its correlation with miR-664-3 p levels in the hippocampus in cerebral ischemia-reperfusion injury(CI/RI)rats.Methods A total of 27 male SD rats were randomly divided into sham operation

CI/RI model and EA groups(n=9 in each group).The CI/RI model was established by occlusion of the middle cerebral artery(MACO)according to the modified Longa methods.EA was applied to"Baihui"(GV20)and"Shenting"(GV24)for 20 min

once a day for 7 days.The Morris water maze tests(place navigation tasks and spatial probe trials)were used to evaluate the rats' learning-memory ability.After EA intervention

the hippocampal tissue was collected to measure the mRNA and miRNA expression profiles by using high-throughput next generation sequencing.Bioinformatics analyses including Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)analyses were performed to screen the most abundant differentially-expressed miRNAs and mRNAs

particularly the target genes of miR-664-3 p.Their biological functional categories including the key biochemical metabolic pathways and signaling pathways involved in CI and EA effect in the left hippocampus tissue were analyzed.Results The average escape latency of place navigation tests was significantly longer in the CI/RI model group than in the sham operation group(P<0.01)

and obviously shorter in the EA group than in the model group(P<0.01);and the times of escape-platform quadrant crossing of the spatial probe trials were considerably fewer in the CI/RI model group than in the sham operation group(P<0.05)

and markedly more in the CI/RI EA group than in the model group(P<0.05).Combined analysis of miRNA and mRNA showed that the differentially-expressed target genes of miR-664-3 p were significantly up-regulated after CI/RI relevant to the sham operation rats(P<0.05)and notably down-regulated in the EA group relevant to the CI/RI model group(P<0.05).The GO analysis displayed that the target genes of miR-664-3 p were significantly enriched in the category of cell transportation activity

ion binding

and transmembrane transporter activity

the production of precursor metabolites and energy

transportation

signal transduction

localization

etc.KEGG analysis showed that the target genes of miR-664-3 p in this study were mainly enriched in the thyroid hormone pathway

Hippo signaling pathway

MAPK signal transduction pathway

cAMP signaling pathway

neurotrophic factor signal transduction pathway

etc.Conclusion EA of GV20 and GV24 can improve the learning-memory ability of CI/RI rats

which is closely related to its effect in down-regulating the levels of hippocampal target genes of miR-664-3 p involving cellular transmembrane transportation

cell to environment interaction

cell-cell communication

etc.