The Long-term Effects Of Dexmedetomidine Pretreatment On Brains Of Neonatal Rats Which Was Anesthetized With Propofol

Objective:To observe the long-sterm effects of dexmedetomidine pretreatment on brains of neonatal rats which was anesthetized with propofol.Methods:Three hundred and ninety seven-day-old（P7）neonatal Sprague-Dawley rats were divided into 13 groups randomly（n=30,each）:group NS,group I,group D₁,group D₂,group P,group DEX,group LY,group TDZD,group PD₂₅,group PD₅₀,group PD₇₅,group LYPD and group TDPD.Rats were intraperitoneally injected（i.p.）with saline,intralipid,or 10%dimethyl sulfoxide（DMSO）in 100μl in group NS,group I,or group D₂,respectively.And rats were intraperitoneally injected（i.p.）with 75μg/kg dexmedetomidine or 1 mg/kg GSK-3βinhibitor TDZD-8 in group DEX or group TDZD,respectively.Besides,5μl 10%DMSO or 5μl PI3K inhibitor LY294002（25μg/5μl）was injected intracerebroventricularly（i.c.v.）respectively in group D₁ or group LY.In group P,rats were intraperitoneally injected with 100mg/kg propofol in the absence of dexmedetomidine pretreatment.25μg/kg,50μg/kg or 75μg/kg dexmedetomidine were administered by intraperitoneal injection respectively 30 min before the exposure to 100 mg/kg propofol in groups PD₂₅,PD₅₀,or PD₇₅.In group LYPD or group TDPD,rats were treated with LY294002（25μg/μl,i.c.v.）or TDZD-8（1 mg/kg i.p.）respectively 30 min before pretreatment with 75μg/kg dexmedetomidine and then propofol exposure.At 2 h after recovery from anesthesia,some rats were used for blood sampling from the left cardiac ventricle for blood gas（n=5）,the others were risen up until 9 weeks old.The spatial learning and memory of rats was testing by Morris water maze（MWM）.The hippocampal neuron apoptosis were determined with TUNEL staining.The expression and cellular localization of PSD95 protein in hippocampus was observed by immunohistochemistry method.The expression of PI3K mRNA,Akt mRNA,GSK-3βmRNA and PSD95 mRNA in hippocampus was detected by Semi-quantitative reverse transcription-PCR.The protein levels of Akt,pAkt（ser473）,GSK-3β,pGSK-3β（ser9）and PSD95 in hippocampus were detected by Western blot.Results:（1）The results showed that the escape latency was longer and the frequency of passing through the platform was decreased in groups P,PD₂₅5 and LYPD as compared with group NS（P<0.05）.Compared with group P,the escape latency was shotter and the requency of passing through the platform was increased in groups PD₅₀,PD₇₅5 and TDPD（P<0.05）.Compared with group PD₇₅,the escape latency was longer and the frequency of passing through the platform was decreased in group LYPD while was opposite in group TDPD（P<0.05）.（2）Compared with group NS,the number of TUNEL-positive cells per mm² in the hippocampal was higher in groups P,PD₂₅5 and LYPD.The number of TUNEL-positive cells was decreased in groups PD₅₀,PD₇₅5 and TDPD compared with group P（P<0.05）.The number of TUNEL-positive cells was increased in group LYPD while was decreased in group TDPD compared with group PD₇₅（P<0.05）.（3）Compared with group NS,PSD95-positive stained neurons were decreased in groups P,PD₂₅5 and LYPD（P<0.05）.Compared with group P,PSD95-positive stained neurons were increased in groups PD₅₀,PD₇₅5 and TDPD（P<0.05）.PSD95-positive stained neurons were redused in group LYPD while increased in group TDPD compared with group PD₇₅（P<0.05）.（4）Compared with group NS,the level of GSK-3βmRNA was increased in groups P,LYPD and LY（P<0.05）,while the level of PSD95 mRNA in group P and group LYPD,the level of PI3K mRNA and Akt mRNA in group LY and group LYPD and the level of GSK-3βmRNA in group TDZD and group TDPD were decreased（P<0.05）.Compared with group P,the level of PSD95 mRNA was increased in groups PD₅₀,PD₇₅5 and TDPD（P<0.05）.However,the level of PI3K mRNA and Akt mRNA was reduced in group LYPD and the level of GSK-3βmRNA was reduced in group TDPD as compared with group P（P<0.05）.Compared with group PD₇₅,the level of Akt mRNA and PSD95 mRNA was decreased while the level of GSK-3βmRNA was inceased in group LYPD（P<0.05）.（5）The results of Western Blot showed that the expression of pAkt（ser473）,pGSK-3β（ser9）and PSD95 was down-regulated in groups P,PD₂₅5 and LYPD as compared with group NS（P<0.05）.Compared with group P,the expression of pAkt（ser473）,pGSK-3β（ser9）and PSD95 was up-regulated in groups PD₅₀,PD₇₅5 and TDPD（P<0.05）.The expression of pAkt（ser473）,pGSK-3β（ser9）and PSD95 was down-regulated in group LYPD while was up-regulated in group TDPD as compared with PD₇₅（P<0.05）.The ratio of pAkt（ser473）/Akt and the ratio of pGSK-3β（ser9）/GSK-3βin groups P,PD₂₅,PD₅₀,PD₇₅5 and LYPD was decreased compared with group NS（P<0.05）.Compared with group P,the ratio of pAkt（ser473）/Akt and the ratio of pGSK-3β（ser9）/GSK-3βin groups PD₇₅and TDPD was significantly increased（P<0.05）.Besides,the ratio of pAkt（ser473）/Akt and the ratio of pGSK-3β（ser9）/GSK-3βin group LYPD were lower,while the ratio of p Akt（ser473）/Akt and the ratio of p GSK-3β（ser9）/GSK-3βin group TDPD were higher than those in group PD₇₅（P<0.05）.Conclusion:Propofol could have long-term effects on brain and damage the spatial learning and memory of in neonatal rats.Dexmedetomidine pretreatment could attenuate propofol-induced neurocognitive impairment,which is mediated by activating PI3K/Akt/GSK-3βsignaling pathway and regulating up the expression of PSD95 protein.