Exploration On Sedation And Hypnosis Mechanism Of YZG-331

NHBA is an adenosine analog isolated from Tianma in the Phytochemical Laboratory of the Institute of Materia Medica,Chinese Academy of Medical Sciences.It has sedative and hypnotic effect,NHBA is transformed into YZG-331 by structural transformation.The previous study of our laboratory found YZG-331 could inhibit the autonomic activity of mice in a dose-dependent manner;significantly prolonged the sleep time of mice induced by pentobarbital sodium,increased the sleep rate of pentobarbital-induced mice,and shorten the sleep latency.Objective:To investigate the role of NMDA receptor endocytosis in the sedative and hypnosis of YZG-331;to detect which sleep awakening nuclei participate in the sedative and hypnotic effects of YZG-331,and to find new targets for YZG-331 sedation and hypnosis.Method:ICR male mice were randomly divided into a solvent control group?15 min group and 60 min group of YZG-331(40 mg·kg-1,i.g.).Western blot was used to detect the protein of CaMKII-NMDAR endocytosis in total cortex,cell membrane and endosome;ICR male mice were randomly divided into solvent control group,1.5h group of YZG-331(40 mg·kg-1,ig),IHC detected changes in c-fos expression in sleep-woke-related nuclei.Results:1.Westem blot analysis showed that the expression of NR1 and NR2B on mouse cortical membrane decreased by 48%(P<0.01)and 23%(P>0.05)15 minutes after YZG-331(40mg·kg-1,i.g.),respectively.The expression of NR1 and NR2B in the cortical endosome increased by 55%(P<0.05)and 67%(P<0.01)15 minutes after YZG-331(40mg·kg-1,i.g.),respectively;The expression of the upstream protein p-NR2B(Ser1303)and its upstream proteins p-CaMKII,which caused receptor endocytosis on the cell membrane,increased by 58%(P<0.05),91%(P<0.01)15 minutes after YZG-331(40mg·kg-1,i.g.),respectively.2.The results showed that YZG-331 increased the expression of c-fos protein in the nucleus accumbens(Acb),ventrolateral anterior region(VLPO),parabrachial nucleus(LPB)by 216%(P<0.01),131%(P<0.01),and 179%(P<0.01)respectively.The expression of c-fos protein in the paraventricular nucleus(PVT)and blue spot(LC)reduced by 50%(P<0.01)and 66%(P<0.01)respectively.In addition,YZG-331 had no significant effect on the expression of c-fos protein in the cortex(cor),the dorsal raphe nucleus(DR),and the dorsolateral(LDT)nuclei.Conclusion:YZG-331 may activate the expression of CaMKII on the cell membrane,then phosphorylate NR2B(Ser1303)to endocytosis of NMDA receptors,reduce the membrane expression of NMDA receptors,attenuate the effects of excitatory neurotransmitters to produce sedative and hypnotic effects;YZG-331 increases the expression of c-fos protein in the nucleus accumbens(Acb),ventral anterior optic anterior region(VLPO)and parabrachial nucleus(LPB),and reduces the expression of c-fos protein in the paraventricular nucleus(PVT)and blue spot(LC);the changes in the c-fos protein expression of these sleep arousal-related nuclei may be related to YZG-331 sedative hypnosis effects,and its specific neuroprojection needs further investigation.