Resuscitative Mechanism Of Borneol And Muscone Based On The Regulation Of Claudins Of Blood-brain Barrier And Their Safety Assessments In Zebraifsh

Objective: This study reports the effect of borneol and muscone on the ultrastructureand Claudins expression of blood brain barrier，to investigate the mechanism of borneoland muscone to promote blood-brain barrier open. Moreover，to assess the safety ofborneol and muscone with zebrafish embryos is the other aim.Methods: After treating adult zebrafish with muscone， the change of BBBultrastructure was observed by transmission electron microscope. The expressionchange of Claudins in BBB at different time points was detected with real-timequantitative PCR technique and whole in situ hybridization.The3hours postfertilization zebrafish embryos were exposed to borneol and muscone at differentculture solutions for96h and inspected daily with microscopy for larval morphology.Spontaneous movements were checked at24hpf. Heart rate at48hpf，hatching rate，embryo deformity rate and mortality rate were evaluated. The expression of sepn1wasdetermined with real-time quantitative PCR technique at96hpf.Results: After administrating muscone，the basement membrane ruptured and wereswollen. The structure of tight junction was loosen while the endothelial cell showedcrimple. And the longer the treating time was，the better the effect was. Borneol ormuscone significantly affected the mRNA levels of Claudin isoforms. Drugs treatmentresulted in a decrease in the abundance of most Claudins，of which Claudin-2，-5a，-7b，-12downregulated significantly.Transcript abundance of Claudin-5a downregulatedmost significantly at1h，and recovered to the normal level at6h with borneol treatment.During4h of muscone treatment，the expression of Claudin-2，-5a，-7b upregulatedtemporarily first，then descent and recovered at last. And Claudin-5a recovered to thenormal level at4h. Borneol and muscone had effect on spontaneous movements，heartrate，hatching rate，embryo deformity rate and mortality rate at different level. At48 hpf，spine curvature，pericardial edema，yolk sac edema，and abnormal swimming wereobserved. Borneol made sepn1gene expression upregulate（P<0.05），while The sepn1gene expression at96hpf in the moscone exposure groups decreased remarkablycompared with that of normal group（p<0.01）.Conclution: Borneol and muscone could make a decrease in the expression ofClaudins，which loosened the tight junction. In addition，the basement membraneruptured and the endothelial cell crimpled. These factors weakened the barrier functionof BBB. Gene reversible downregulation demonstrated that the open of BBB wasphysiological and the change would not bring about brain impairment. From the clinicalsafe medication perspective, borneol and muscone at high concentrations had significanttoxic effects on the development of zebrafish embryos.