Neurochemical Mechanisms Of YZG-331 On Sedative-hypnotic Effect

YZG-331 is a new compound derived from the gastrodia elata with sedative and hypnotic activity.The structure is novel.It is proposed to apply for class 1.1 new drugs of chemical drugs.Biochemical analysis is one of the common methods to study the pharmacological mechanism of sedative-hypnotic drugs in different parts of brain,especially on neurotransmitters.Therefore,this study explored the sedative-hypnosis mechanism of YZG-331 from the perspective of neurochemistry.Preliminary experiments showed that YZG-331 could dose-dependently inhibit the autonomous activities of animals,and dose-dependently extend the NREM sleep time of mice,shorten the sleep latency time,and have a significant sedative-hypnotic effect.In the combined experiment with superthreshold dose of pentobarbital sodium,the sleep prolongation rates of YZG-331(5,10,20 mg/kg,i.g.)on mice were 23.7%(p=0.09),46.2%(p<0.05),and 82.8%(p<0.01),respectively.YZG-331(5,10,20 mg/kg,i.g.)could significantly shorten the latency for mice to fall asleep.The results were consistent with previous studies in laboratory,proving that the efficacy of YZG-331 on sedative-hypnotic was significant and stable.On these basics,the next parts were to study its mechanism.In these experiments,HPLC was used to detect the effect of YZG-331 on the contents of neurotransmitters in the microdialysate and homogenate of the dorsal raphe nucleus(DRN),dorsal striatum(CPu),external globus pallidum(GPe)and orbitofrontal cortex(OFC).The results of microdialysis experiment showed that YZG-331(40 mg/kg,i.g.)could significantly increase the microdialysate GABA level in the DRN,CPu,GPe,and OFC of mice(DRN 104.1%,p<0.05;CPu 70.8%,p<0.01;GPe 59.5%,p<0.01;OFC 20.8%,p<0.01).The level of Glu was only significantly increased in the CPu(19.8%,p<0.01).There was no significant changes in NE content in the measured nuclei.The content of DA decreased significantly in the CPu at 60-90 min after administration(31.6%,p<0.01),but did not change significantly in the DRN.And the DA content in GPe and OFC dialysates were lower than the detection limit.The neurotransmitters in homogenate results showed that YZG-331(40 mg/kg,i.g.)had no significant effect on the content of GABA and Glu in GPe.The content of DA in CPu decreased after administration YZG-331,but with no significance;DOPAC content was not significantly changed.There was no significant change in NE concentration in the tissue homogenate of prefrontal cortex and hypothalamus.The increase of NO content in the brain could induce animals to fall asleep.The results showed that YZG-331(40 mg/kg,i.g.)could significantly increase the content of NO in the brain of mice(26.9%,p<0.01).The results of NOS activity detection showed that YZG-331(40 mg/kg,i.g.)could increase the activity of cNOS in the brain of mice(34.6%,p=0.064,n=3),but had no effect on the activity of TNOS and iNOS.Western Blot results showed after administration YZG-331(40 mg/kg,i.g.),the expression of p-CaMKⅡ was significantly reduced in hypothalamus and DRN.In conclusion,the results of this study found that YZG-331could significantly increase the GABA content in the microdialysate of CPu,GPe,OFC and DRN,enhance the inhibitory effect mediated by its binding to GABAa receptor,and decreased the activity of neurons in the brain,which may be one of the sedative-hypnotic mechanisms of YZG-331.YZG-331 had no significant effects on NE content in both of extracellular fluid and homogenate in various brain regions of mice,suggesting that NE did not take part in the sedative-hypnotic mechanism of YZG-331.YZG-331(40 mg/kg,i.g.)also could inhibit CaMKⅡ posphorylation in mice hypothalamus and DRN,increase nNOS activity of brain,which may lead to the raise of NO content in brain.It has been reported that the increase of NO content could induce the increase of adenosine in basal forebrain or s-nitrosylation protein disulfide inomerase led to the orexin peptide incorrect folding and inhibiting neurons activity to promote sleep.This may be another mechanism of YZG-331 sedative-hypnotic effect.